Light Antennas in Phototactic Algae

59 Pages·2007·10.08 MB·English

MICROBIOLOGICAL REVIEWS, Dec. 1980,p.572-630 Vol.44,No.4
0146-0749/80/04-0572/59$02.00/0
Light Antennas in Phototactic Algae
KENNETH W.,OSTER'* AND ROBERT D. SMYTH2
DepartmentofPharmacology,MountSinaiSchoolofMedicineofthe ty UniversityofNew York, New
York, New York 10029'; andDivisionofBiology, CaliforniaInstituteofTechnology, Pasadena, California
911252
INTRODUCTION ..............6.............573
TheProblem: DetectingLightDirectionintheNativeEnvironment ..... 574
....
The Solution: aDirectional AntennaDesignedto Match theEnvironment of
theOrganism .......................................................... 576
ANTENNASTRUCTURES 579
....................................................
OpticalPrinciples 580
Absorption 581
...............................................................
Scattering 581
Refraction 581
Reflectionandinterference 581
...............................................
Waveguideoptics ........................................................ 583
Dichroism 584
................................................................
TrackingAntennaDesigns 584
..................................................
Chlorophyceae: multilayerquarter-wave stackantennas .................. 584
Chlorophyceae: variationsindesign .... 588
..................................
Dinophyceae: quarter-wave stackantennasandlensantennas 590
...........
Cryptophyceae: dielectricslabwaveguideantennas 592
......................
Euglenophyceae: absorbingscreensanddichroiccrystaldetectors 592
.......
Chrysophyceae,Xanthophyceae,andPhaeophyceae:absorbingscreensand
paraflageliarswelling 593
.................................................
Eustigmatophyceae: absorbingscreensandparaflagellarbuttons 594
........
Prymnesiophyceae(Haptophyceae) 595
.......................................
Discussion 595
ANTENNAPROPERTIESDETERMINEDFROMBEHAVIOR 596
.................
Threshold: ReceptorPigmentConcentration 596
Receptorpigmentcontentofvariousalgae 598
.....
Action Spectra: Identity ofPhotoreceptor Pigment andMechanisms ofPro-
ducingDirectivity 600
....................................................
Thresholdactionspectrum 601
...............................................
Finite-responseactionspectra 601
............................................
Algaewithlayeredeyespots 601
......
(i) Volvox ............................................................. 601
(ii) Chlamydomonas 602
(iii) Platymon as 602
........................................................
Dinoflagellates ... ..... 603
...
Cryptomonas 603
.............................................................
Euglena 604
..............................................................
Intensity-ResponseCurve:IdentityofReceptorPigment,MechanismsofPro-
ducingDirectivity, andPigmentRegenerationRate 604
...................
Chlamydomonasactionspectraandregenerationrate 606
....................
Polarized Light: Characterization of the Dichroic Crystal Detector in
Euglena 607
DiscussionofReceptorPigments ........... 608
... ...
ANTENNAFUNMCIONINPHOTOTAXIS 609
........
SignalProduction: ScanningbyCellMotion 609
.. ....
SignalProcessing 6............12
Response:TrackingtheLightDirection 6............................6.1.
Chknmydomonas ........................ 616
..
Microthamniun 617
Eugklna ........... 617
........... .. .. ..
Dinoflageliates ..................................... 617
Colonialalgae .. 617
... ........ .. .. .....
Discussion 619
PROSPECTSANDCONCLUSIONS 619
...........................
572
VOL. 44,1980 ALGAL PHOTOTAXIS 573
APPENDIX ......................................... 621
AnalysisofIntensity-ResponseCurves....................................... 621
Independenteffectofphotoreceptorandscreen .......................... 621
Derivationfromreceptorkinetics ......... 621
LITERATURECITED 623
.........................................................
It's my experience that everyone who has a cell drifted away. By means ofconverging and
device like a tracker in mind wants to use it diverging lenses, Buderproved that the cell re-
... where the range is maximum, the signals sponds to light direction, not an external gra-
aresmall, andthegoingistough. dient oflight intensity. In other words, a posi-
LucienM. Bibernan (19) tively phototactic cell swims toward a light
source whether the extemal light intensity in-
INTRODUCTION creases, decreases, or remains the same. Buder
All photosynthetic organi8ms have evolved examined the swimming paths and aggregation
specializedmeansofregulatingtheirexposureto patterns of a variety of algae, including
sunlight. One form this adaptation has taken Chlamydomonas, Carteria, Euglena, Trache-
among the flagellated algae istheirphototactic lomonas, colonialalgae,andswarm cellsoffila-
response, the ability to swim toward or away mentousforms.Hefoundthatthesealgaeorient
from a source of light. The response is found to steady illumination and concluded that this
among representatives ofall ofthe major algal behavior is the biologically gnificant photore-
groups except the Cyanophyta and the Rhodo- sponse.Inshort,flagellatedalgaearespecialized
phyceae, which entirely lack flagellated forms. fortracking lightdirection.
(We use the provisional classification ofDodge The geometry ofboth the cell and its swim-
[58] in which most groups are not referred to ming path are important for phototaxis. When
above the level ofclass because ofpresent un- the cell changesitsorientation to light, the dis-
certainties of relationships. This classification tribution of light within the cell changes. Jen-
followsbotanicalratherthanzoologicalnomen- nings (131) and Buder (31) recognized that the
clature.) The response was first observed by cellusesthis change as a signal ofhowthe cell
Treviranus in 1817 in zoospores released from has deviated from the light direction and what
thefilamentousalgaDraparnaldia (239).Lead- correctiveresponseisneeded.However,detailed
ing biologists studied the response in the nine-
teenthandearlytwentiethcenturies,aperiodof
investigation thatculminated inBuder'sdefini- B B
tivepaperonalgalphototaxisin1917(31).After
aperiodofrelativeneglect,phototaxishasbeen
under active investigation in a number oflabo-
ratories in recent years. Earlier work is thor- A A
oughly reviewed by Haupt (106); later work is
covered by Feinleib and Curry (75), Forward
(80), Lenci and Colombetti (146), Diehn (54),
and Nultsch and Hader (200). Here, we apply
physical principles to obtain new information
from the results of physiological experiments
and ultrastructural studies ofalgae reported in
the literature. Thepurpose ofouranalysisisto
explain phototaxis better and to suggest new
approachesforstudyingtheproblem. FIG. 1. Stereoplotofthepathofanalgashowing
Buder (31) demonstrated thatphototactic al- phototactic attraction to light. Thepath appears in
gal flagellates orient to the direction ofa beam correctthree-dimensionalperspectivewhentheimage
oflight, provided its intensity is within the be- ofthe right-handplot seen by right eye is superim-
havioral range ofthe cell. (We use the familiar posed on image ofleft-handplot seen by left eye. A
termlightintensity. More and more frequently single ceU of Chlamydomonas reinhardtii (strain
irradiance is used instead.) This behavior is il- 137c, mating type +) was followedfor 12.0 s in the
trackingmicroscope(16).Dimredlight,whichcauses
lustrated in Fig. 1, which shows the swimming
no response, was usedfortracking. Theposition of
path ofa single cell. At B, light was turned on thecellwasrecordedevery42ms. Thetrackstartsat
from below. The cell altered itspath, locked on A.AtB,bluestimulatinglightfrombelowwasturned
to the beam, and swam directly toward its on. At C, thestimulating light was turned off. Path
source. At C, the light was turned off, and the length, 1.9mm;averagespeed, 159± 14pun/s.
574 FOSTER AND SMYTH MICROBIOL. REV.
information about the signal and the response bacteria have overcome this problem to some
hasproveddifficulttoobtain. Afterconsidering extent by evolving rod and spiral shapes. Even
phototaxisfromasmanyviewpointsaspossible, so, bacteria show at most a limited ability to
we concluded that specialized structures inde- orientparallel orperpendicularto alightbeam
pendent of the photosynthetic apparatus are (K. W. Foster and H. C. Berg, manuscript in
used for phototaxis, and that these structures preparation). They cannot sense lightdirection
are best thought of as directional light wave and cannot track. For most phototactic algae,
antennas. We call them antennas because, like the increase in size makes rotational Brownian
radio wave antennas, their optical characteris- motion insignificant. For example, Brownian
tics are dominated by interference and diffrac- motion randomizes the orientation of a sphere
tion. In both algal and radio antennas this is a 10umindiameter,acommonsizeforunicellular
consequence oftheir size, which is ofthe same algae,inabout6.5min (relaxationtime). Thisis
orderastheelectromagneticwavestheyreceive. much longer than the reaction time ofthe cell.
In both cases the size of the structures is less Furthermore, internal gradients of light inten-
than50wavelengths. (Largerradioantennasare sity can be created in a surprsing number of
properly calledtelescopes and can bedescribed ways (see Antenna Structures). (The behavior
infamiliar opticalterms.) of the smaller phototactic algae has not been
Usually, an algal antenna is an eyespot plus studied in detail. The smallest is Micromonas
associated structures. Algae controlphototactic pusilla,amonoflagellate 1to3,umlongand0.75
behavior by scanning their environment with to 1.0pinwidewithavolumeof1 to2pm3[158,
the antennas. In our discussion, we follow a 164, 237]. Like bacteria, this orgaiism swims
philosophyusedindeterminingperformancecri- rapidly [50 body lengths per s] and changes
teriaofman-made antennas, inwhichemphasis directionfrequently.)
isonoptimumdesignforsomeintendedfunction Tracking is a sophisticated form ofbehavior
(224). In biological terms, this means that we involving problems of timing, stability, and
must consider the environment, the signal re- noise. Acquisition of this ability required the
ceived by the photoreceptor, signal processing, evolution of complex cellular machinery and
andtheresponseinadditiontothepropertiesof integration of the specialized components into
the photoreceptor structure. Relevant informa- an effective design. Tracking cannot be the in-
tiononphototaxisislimited, butbyintegrating cidentalby-productofresponsesprimarilyused
detailsfrom avariety ofalgae, comparing them for other purposes. One biological purpose of
with other organisms, and considering the re- phototaxisistomovetheorganismasrapidlyas
quirements for signal processing, we can map possible into the proper intensity of sunlight.
out the general principles of algal phototactic The cellsmay also orientto optimize theirrate
behavior. of photosynthesis. Some algae appear to use
One ofouraimsistoprovide thereaderwith lightdirection tofind aplaceto mateorgermi-
anintuitive sense ofhowphototaxisworks. For nate. Oneindicationthatalgaeareunderstrong
thispurposecertaindimensionsmustbekeptin selectivepressuretodevelopphototacticbehav-
mind. Typical swimming speeds are 100 um/s ioristhespectacular convergentevolutionthat
for unicellular algae and up to 1,000 pim/s for hasoccurredindifferentalgalgroups.Aswewill
large colonial algae. Swimming iscausedbythe see, many examples are known of diverse ele-
flagellar beat. Asthe organismsswimn, theircell ments that have been adapted to serve much
bodies rotate at frequencies in the range 0.2 to thesame functionbydifferentalgae.
2.0Hz. Theflagellarbeatfrequencyis 10to 100
times greaterthantherotationfrequency. Pho-
TheProblem: DetectingLightDirectionin
totactic algae range from the size ofbacteria to
theNativeEnvironment
colonial algae containingthousands ofcellsand
having a diameter up to 1 mm. Bacteria are We have seen that algaeregulate their expo-
typically afewmicrometers inlength. suretolightbyswimmingtowardorawayfrom
This increased size relative to bacteria is im- thelightsource.Todothistheydetectapattern
portant to the way that flagellated algae carry in the distribution oflight reaching them. The
out phototaxis. Bacteria are too small to track pointisobviouswhenwe considerthatorienta-
light. One reason for this is that they cannot tion would be impossible ifthe light were uni-
create within their bodies a sufficient gradient formly distributed. Light in natural environ-
oflightintensityto obtainaccurate cluesabout ments has three physical properties (namely,
light direction from changes in light intensity color, polarization, and intensity) whose distri-
caused by their own motion. Furthernore, the bution pattem could be used. There is no evi-
swimming direction of bacteria is rapidly ran- dence thatalgaeorientbyeithercolororpolar-
domizedbyrotationalBrownianmotion. Motile ization, eventhough theyhavesomesensitivity
VoL 44,1980 ALGAL PHOTOTAXIS 575
to both qualities. They orient to the intensity ration]. The flagellar beat frequency has been
pattern. measured at 25 Hz [125] and 50 Hz [227] with
Thesimplestpossiblepatternisparallellight, different conditions andstrains. In one beat cy-
butnatural light, especially under water, is dif- cle, rotational diffusion causes the cell to turn
fuse, coming from different directions. The or- an average of80. The swimming speed is typi-
ganism therefore must orient toward the aver- cally 100,um/s. Wehaveinourpossessionhigh-
age,orresultant,lightdirection.Wearespeaking speed motion pictures of swimming Chlamy-
nowoftheintensityoflightatasinglepointand domonas cells. These filns, made in the labo-
howitisdistributedasafunctionoftheangleof ratoryofT.L. JahnbyJ. F. Fonseca, showthat
incidence. We are notspeaking ofa spatialgra- duringonebeatcyclethecellgoesforwardabout
dientoflightintensity. Thedistribution oflight 4jmandbackwardabout2pm.Duringonebeat
intensitymustbesensedinsuchawaythatthe cycle the cell may turn back andforth through
organismreceivesasignaltellingithowtomake anangle ofup to 20 or300.) (ii) A second noise
a precise correction, a correction of the right source isconvection currents. In shallow water,
magnitude in the right direction. For greatest heating on the bottom causes water to rise to
effectiveness, the organism should respond cor- the surface. This creates a series of circulation
rectlyfrom anyorientation. pathsthatcontinuouslycyclethewaterfromthe
Foursevere constraints areplaced onthe de- bottom to the surface and back to the bottom
signofthelightdetector: itmustfindthesignal again.Thealgaemustbeabletoreorientasthis
in a background ofnoise, it must operate over convective flow carries them off course. Both
mostoftherangeoflightintensitiesfoundinthe convectioncurrentsandrotationaldiffusiondif-
native environments, it must have the proper fer from the following sources of noise in that
spectralsensitivitytodetectthewavelengths of the signal is altered by actual variation in the
light that reach the algae, and it must commu- orientation ofthe organism to the light. (iii) A
nicate rapidly to the response apparatus. The thirdnoisesourceisexternalfluctuationinlight
design ofa detector that extracts useful signals intensity. Waves on the surface ofany body of
from noise is complex and calls for a detailed wateractaslensesthatpartlyfocuslightbelow
discussionhere ofthenoise problem. the surface. As the waves move, bright bars of
Atleastfive sources ofnoise seem important light move with them, causing the light below
in phototaxis. (i) Rotational diffusion is one the surface to fluctuate. The phenomenon can
source ofnoise. As already discussed, algae are beseenbylookingatripplesinshallowwaterin
solargethatrotationalBrownianmotion,which directsunlight. Measurementsmadeusinglarge
resultsfromthermalbombardmentoftheorgan- sensorsinlargebodiesofwaterexposedtodirect
ism bywater molecules, is insignificant. This is sunlight (51, 195 and references therein, 225)
not the only cause ofrotational diffusion. The indicate that the fluctuations are large. For ex-
motility of the organism itself has a random ample, in the upper few meters the variation
componentthatconstantlyaltersitsorientation. may be as much as 80 to 90% of the mean
Repetitive beating oftheflagella moves the or- intensityandismaximuminthefrequencyrange
ganism.Ifallflagellarbeatcycleswereidentical, 0.4 to 1.0 Hz. Thisis somewhat below the rota-
the organism would be driven along a uniform tion frequency of unicellular algae. Measure-
helical path (153) perturbed only by the other ments with sensors the size ofalgal sensors are
sourcesofnoise. Thisispartlywhatoccurs, but needed ofthe magnitude oflight fluctuation as
random variation from one beat to the next a function of frequency in algal environments.
introducesrotationalandtranslationaldiffusion. We predict that appropriate measurement will
The effect is accentuated by the low Reynolds show thatexternal fluctuation inlightintensity
number associated with cell motion; that is, in- constitutes an important source of noise. (iv)
ertial forces are negligible compared with vis- Shotnoiseiscausedbystatisticalfluctuationin
cous forces. Thishasthe consequence oftrans- thenumberofexcitations ofphotoreceptorpig-
mittingeverymotionoftheflagellatomotionof ment,anditsmagnitudedependsontheamount
thebody,causingittomovebackandforthwith ofpigmentandtheamountoflight. (v) Internal
each flagellar beat. In Chlamydomonas, where noise is noise produced within the organism by
it has been measured, rotational diffusion is such factors as concentration fluctuations and
greatenoughsothatcoursecorrectionsarecon- fluctuations in membrane potential. This noise
tinuouslyrequired,butitdoesnotpreventtrack- occurs at all stages, from signal reception
ing. (Chlamydomonas has a Reynolds number through response. We know from the relatively
of10-3. Itrotatesinaleft-handhelixatabout2 smallnumberofmoleculesinvolvedinbiochem-
Hz. Superimposed upon the rotation is rota- ical processes that these fluctuations must be
tionaldiffusion witharelaxation time of2 s [R. large and must therefore introduce appreciable
D. SmythandH. C. Berg, manuscript inprepa- noiseintophototaxis, butatpresentwehaveno
576 FOSTER AND SMYTH MICROBIOL. REV.
adequate measure or good theoretical estimate Environmental
ofthisnoise.
Light Pattern
The Solution: aDirectionalAntenna
DesignedtoMatchtheEnvironment ofthe
\; 11~i
Organism
Phototacticalgaesolvetheproblemoffinding
the light direction by scanning their environ-
Flagella
ment with an antenna sensitive to light. The
antenna must be integrated with the response 70(
mechanismofthecellsoastoproduceaneffec-
tive overall design. Consider unicellular forms
first. Despite their enormous diversity ofform,
theyuse the same basic scheme forphototaxis.
The extent to which their antenna design
matches requirements determines how effec-
tivelytheytrack.
Figure2showsadiagramofChiamydomonas
thatillustrates the principle. The cell hasared
spot on its side, the eyespot. The eyespot and
associatedstructuresformadirectionalantenna
(seeAntennaStructures). (Mostofthealgaewe
discuss have eyespots. This means that one of
the components of the antenna is visibly pig-
mented.However,thevisiblepigmentisnotthe
photoreceptor pigment, which is present in too
small an amount to be seen in the microscope.
Itisnotessentialthattheantennahaveavisibly
pigmented structure, and algae exist that have
antennas but not eyespots. We use the words
eyespot and stigma synonymously.) The an-
tenna senseslight moststrongly when thelight
is normal to the outer surface of the antenna
and least when it strikes the antenna from the
sideortheback. Thecellswimswithitsflagella
forward, rotating approximately twice a second
asitswims. Thisrotationcausestheantennato
scan the environment. If the swimnming path Antenna
were held parallel to the light direction, the b. \direction
antenna would receive nearly constant illumi-
nation throughout the scan cycle. But noise FIG. 2. Design principles ofphototaxis in Chla-
mydomonas. (a) Side view ofcell; (b) endview. The
quickly causes the cell to deviate from the
incident lightpattern is indicated by solid arrows.
aligned orientation. When the swimming path
Theeyespot,whichliesinsidethechloroplast(dashed
becomes sufficiently inclined to the light direc- line), formspartofthe antenna. Rotation ofthecell
tion, the light intensity seen by the antenna causes the antenna to scan the incident light. This
becomes periodic. This results from the varia- producesasignalthatcontrolstheflagellarbeat(see
tioninanglebetweentheantennaandthelight Fig.3). Theantennadirection(openarrow)isnormal
direction that occurs during the rotation cycle. to the cell surface. The antenna ismostsensitive to
The cell processes thisperiodic signal to deter- lightcomingfromthisdirection.Successivepositions
mine when to make a corrective response that ofthe flagella during thepower stroke are shown.
FlageUarmotioncausesthecelltotranslatewiththe
will realign the path to the light. The response
flagellarendforwardandto rotate in the left-hand
of the flagella, by changing the orientation of
sense.
the cell, changes the shape ofthe signal during
thenextscancycle.Thesequenceofeventsthus thatthecellreceivesapatternedsignaliscentral
forms a closed loop, the tracking servo loop to phototaxis. The pattern is used in at least
diagrammed inFig. 3. three important ways: for the infornation it
We thus see that rotation ofthe cell gives a contains about orientation, for noise suppres-
temporalpatterntothelightintensitysensedby sion, andforextension ofthe effectiveintensity
the antenna (see Signal Production). The fact range. We describe each of these uses briefly
VOL. 44,1980 ALGAL PHOTOTAXIS 577
Intensity at cell
Antenna signal
I rce> r v
\1
Processed signal
FIG. 3. Control loop for phototaxis showing environmental stimulus and the signal at two stages of
processing. The signal intensity (ordinate) during two rotations of the antenna is plotted against time
(abscissa). Thesignaldrifts becauseofnoise, buttheprocessedsignalhastwowell-definedmaximaphased
withtherotationofthecell.Signalswerecomputersimulatedwithoutattemptingrealisticnoisesimulation.
here,andanalyzethemindetailwhenwediscuss sensor asymmetrically located in the cell pro-
antennafunction (see Signal Processing). duces some directionality. Increased direction-
Alltracking devicesworkbyprocessingsome ality is achieved by specialized antenna struc-
form oferror signal that is generated when the tures which employ a variety of optical princi-
trackerdriftsofftarget.Tobeusefulfortracking, ples (see Antenna Structures). This increases
theerrorsignalmustcontaininformationabout the amplitude of the signal with concomitant
the direction ofdeviation from the target. The improvementintracking.
quality oftracking can be improved ifthe error Whatpattern ofdirectionalityinthe antenna
signalalsocontainsinformation aboutthemag- produces the maximum amplitude of signal?
nitude of the deviation. Probably the most fa- Qualitatively, we can see that a narrow direc-
miliar kind of tracker is a man-made one in tionalitywouldmakeitdifficultfortheantenna
which an array of sensors points toward the to track a broad source, because the antenna
source being tracked. Each sensor points in a would wander around on the source without
slightly different direction. When the tracker detectingmuchchangeinintensity.Bythesame
driftsofftarget,thepatternofstimulationofthe reasoning, a wide antenna would be poor for
sensors changes; the signal processor then ana- tracking apoint source. The optimum designis
lyzes the pattern of change and figures out in tomatchthedirectionalityoftheantennatothe
what direction and how fast it must move the distribution ofthe light it tracks. For the ideal
tracker to return it to the target. Unicellular tracker, in other words, a plot of the relative
algae have onlyonesensor (seeAntenna Struc- sensitivityoftheantennainalldirectionswould
tures). In a single-sensor tracker it would be becongruentwithaplotofthelightintensityin
uselesstopointthesensoratthetarget,because all directions. We would predict, therefore, a
thenthesignalchangeproducedupondeviation rough match between the directionality ofthe
would contain no directional information. In- antennaandthelightdistributionthatitismost
stead, the sensor must point offto the side of advantageous for the organism to detect (see
the target, anditsdirection must bevariedsys- Signal Production).
tematically to produce a periodic signal. The The second use that the cell makes of the
phase ofthe signal contains information about temporalpatternofstimulationisnoisesuppres-
the error direction, and its amplitude contains sion. Findinganappropriatesignalinthemidst
information aboutthe errormagnitude. Theer- ofnoise is never a trivial task. The problem is
rorinformationcanbeusedbythecellifthecell simplifiedwhenthedetectorcantakeadvantage
makes a fixed response that is phased with the of some known characteristic of the signal. A
errorsignalandproportionalinmagnitudetoits familiar example is AM radio, in which the re-
amplitude. ceiver can be tuned to the frequency of the
For the rotation ofthe cell to produce a use- transmittingstation. Evenifthe signaldoesnot
fullypatternedtemporalsignalatthesensor,the alreadyhaveausefulcharacteristictoaiddetec-
antenna must be directional. By this we mean tion when it arrives, such a characteristic can
that the intensity sensed by the antenna must sometimes be imparted to it before it reaches
vary with the angle of incidence of the light the sensor. This is the scheme employed by
reaching the antenna. Merely having a small phototactic algae. They use their own rotation
578 FOSTER AND SMYTH MICROBIOL. REV.
togivethelightreceivedbythesensoraknown characteristics, namely,thesignalcausedbythe
temporal pattern that can be matched with a flagellar beat. Later we discuss how this might
signalprocessor (see Signal Production). be used in signal detection [see Signal Process-
When considering noise, it is convenient to ing].)
thinkoftheinputstimulusinawaythatasignal We can ask why evolution has selected the
iscommonlyanalyzed inelectronics, namely, as particular rotation frequencies observed. They
the sum of a series of sine waves of different probably represent, at least partly, a compro-
frequencies. The amplitudes of the sine waves mise with the frequency spectrum ofthe noise.
are different atdifferent frequencies and, taken The longer the rotation time, the more accu-
together, determine the form ofthe inputstim- rately the signal can be known. A limit to the
ulus. From this point of view we can say that length of the rotation period is set by low-fre-
cell rotation greatly enriches the sensed light quency noise, in particular noise caused by ex-
intensity for certain frequencies. The signal ternal fluctuation in light intensity and rota-
processorthenactsasaband-passfiltertoselect tional diffusion. Rotational diffusion eventually
those frequencies andsuppress others. rendersoldinformationinvalid. Wesuspectthat
Thefilterusedbythesignalprocessorismore noise from these souces rises at frequencies
thanjustaband-passfilter. We showlaterthat lowerthantheobservedrotationfrequency,and
itspreciseform canbefound experimentallyby thattherotationfrequencyhasbeensetcloseto
measuringtheimpulseresponse,thetimecourse the lowest frequency where phototaxis is possi-
of response after a pulse stimulus (see Signal ble.
Processing). The filter performs two functions: Theschemeofsignalmodulationandprocess-
(i) signalmatchingand (ii) differentiation. ing described so far is precisely that which has
Bysignalmatching,wemeanthatthedetector been demonstrated in the phototropic response
is especially sensitive to a particular shape of ofthefungusPhycomyces (50,82), althoughthe
inputsignal,notmerelytoaparticularfrequency time scale for algal phototaxis is shorter by a
range. One expects a rough match between the factorof103.
processingfilterandsomestimuluspatternthat The third use mentioned above for a pat-
can be produced by the rotation ofthe cell and terned signal is range extension. We have seen
bythe directional properties ofthe antenna. that the processor acts as a differentiator. Be-
Differentiation is important in suppressing sides suppressing low-frequency noise, differen-
low-frequency noise. Low-frequency noise is tiationalsomakestheprocessedsignalrelatively
troublesomebecauseitrepresentsashiftingbase independent ofthe average intensity ofthe in-
line uponwhich the signal issuperimposed and cident light. The intensity range for algal pho-
renders absolute measurements meaningless. totaxisisbestdocumentedforChlamydomonas
Therateofchangeofthesignal (thederivative) (76, 201), which is phototactic over a 104 range
ishighly insensitive to thisform ofnoise. Many of intensity extending from 1011 to 10'5 quanta
biologicalsensorstherefore measurethe deriva- cm-2 s-I at 500nm.
tiveratherthanthe absolute levelofthe signal. This operating range must match the range
Some well-studied examples are bacterial che- relevant to the survival of the organism in its
motaxis (30), the light-growth response ofPhy- natural environment. Its lower limit is set by
comyces (82, 150), the electroreceptors of the several factors, including the directionality of
ampullae of Lorenzini in dogfish (192), some theantennaandthelengthoftimetheprocessor
mechanoreceptors in skin (33), and carotid bar- integrates the signal (see Threshold). Because
oreceptors (204). In algae, the antenna signal is the variation in these parameters is limited,
alsoprocessedtodetecttherateofchangeofthe threshold is primarily determined by the total
signal. amount ofeffective receptor pigment. This fol-
In receptors, high-frequency noise (frequen- lowsfromthefactthattheprobabilityofphoton
cies above the band-pass region) is suppressed captureisdirectlyproportionaltotheamountof
by averaging the input stimulus over a time receptorpigment inthe cell: themore pigment,
longer than the period of the high-frequency the greater the possible operating range. Be-
noise. In other words, the signal is integrated cause of the quantal nature of photons, this
over the memory time ofthe receptor. Applied lowerlimitisabsoluteandcannotbeloweredby
to algae, this means that the response at any any amount of processing or amplification. In
time is determined by information collected principle, a photoreceptor could operate up to
throughout the rotation cycle. This suppresses very high light intensities, but usually it is ad-
noise, but creates a delay in the response that vantageous to sacrifice performance at high in-
must be compensated for in the design (see tensityforoptimumperformance atsomeinter-
Signal Processing). (The input signal possesses mediate intensity.
one high-frequency component with known Phototaxisalsorequiresthatthespectralsen-
VOL. 44,1980 ALGAL PHOTOTAXIS 579
sitivitymatchthecoloroflightthecellcanmost ray.Thefunctionofthearraycanbeinterpreted
advantageously use to control its response (see from an understanding of antenna function in
Action Spectra). Both the photoreceptor pig- unicellularforms.
ment and the antenna structures that interact Wedonotdiscussoneofthemostfundamen-
with the light must be effective in the same tal aspects of phototaxis, namely, the factors
spectralrange.Furthermore,thechoice (aswith that determine whether a cell goes toward the
photosynthetic pigments) is limited to visible light or away from it. We do not know what
light, the only light appreciably transmitted by design features favor positive or negative pho-
water. The photosynthetic pigments, which ac- totaxisorhowtheindividualorganismregulates
count for most of the absorption of photosyn- whichwayitgoes. Weassumethatinsomeway
thetic cells, absorb stronglyinthe blue andred this has already been detennined, and we ana-
ranges; there is a range of intermediate wave- lyze the tracking technique used. The tracking
lengths where their absorption is weaker. In principle is the same for either positive or neg-
many phototactic algae, at least part of the ativephototaxis.
phototactic range lies at these intermediate Finally, we donotconsiderin detailwhatthe
wavelengths.Thisallowsthealgaetorespondto flagellum doeswhenitrespondsto lightorhow
light evenwhenshaded by overlying photosyn- itsresponse affectsthemotionofthe cell. Fora
theticorganisms. comprehensive review of these subjects, see
Finally,theproblemofrapidandfaithfulcom- Jahn and Bovee (126-128). Alteration offlagel-
municationofthesignalfromtheantennatothe larshapebylightanditseffectonthe direction
flagellumiscritical.Ifthereceptorweretocover of flagellar response have been described by,
alargeportionofthebody,thenthesignalcould amongothers,Metzner(185),HandandSchmidt
be spread out in time due to different delays (105), and Diehn et al. (55). Throughout our
from different parts ofthe receptor. Such tem- discussion, we assume that the direction ofre-
poralspreading would decrease the directional- sponseisfixedwithrespecttothecell, andthat
ityoftheantenna. Thesolution tothisproblem thetimingoftheresponseiscriticalfortracking.
isasmallantennaand extremelyrapidcommu- Thisintroductorydiscussionofphototaxishas
nication, less than 20 ms in Chlamydomonas introduced severalprinciplesthatmaybe unfa-
(see Response). Forsuchrapidcommunication, miliar.Theyshouldbecomecleareraswediscuss
largedistancesmustbecoveredelectrically; dif- available information on ultrastructure, behav-
fusion can be used only for short distances or ior, andfunction.
wherepoorperformanceistolerated.Wediscuss
ANTENNA STRUCTURES
below various ways that algae have found to
meetthisproblem (seeAntennaStructures, Re- In this section we examine the ultrastructure
sponse). ofthe antennas ofdifferent algae to determine
Atthebeginningofthisintroductionweposed thedistributionoflightwithinthestructures. In
the questionofhowamicroorganism solvesthe particularweconsiderhowthelightdistribution
formidable problem ofdetectinglight direction. varieswiththeorientationofthecellrelativeto
We note that it requires a series of matches the light direction. If we know the location of
betweentheorganismanditsenvironment.The the photoreceptor, we can then ask what the
directionalityoftheantennamatchesthespatial directional sensitivity of the antennas is. In
lightdistribution.Therotationfrequencyofthe keeping with antenna terminology (41), we use
organismmatchesafrequencyatwhichtheratio directivityasameasureoftherelativesensitivity
ofsignaltonoiseisoptimum.Thetemporalfilter of the antenna in a particular direction. For a
of the signal processor matches the temporal strictdefinitionofdirectivity,imaginethatatest
variation ofthe scanningsignal. The functional beam of parallel light is shone at the antenna
rangeofthe detectormatchestherangeofinci- from a particular direction, and that the inten-
dent light intensity. The spectral sensitivity of sity received by the sensor of the antenna is
the antenna matches the color ofincident light measured. The directivity is this intensity di-
most useful for controlling behavior. The an- vided by an isotropic reference, namely, the
tenna must be integrated with the response average intensity received by the sensor when
mechanismofthe cellforrapid communication the test beam is moved through all angles of
ofthesignalandproperphasingoftheresponse. incidence. Thedirectivity isproportionalto the
Muchofthefascinationofphototaxisconsistsof ratio ofincident to received light intensity in a
tryingtounravelthiscomplexinterplaybetween particulardirection.Iftheantennahadnodirec-
theorganismanditssurroundings. tionalsensitivity, the directivitywouldbe 1.0in
Wediscusscolonialalgaebelow (seeAntenna all directions. Normally it is greater than 1.0 in
Function). Note,however, thatinstead ofasin- some directions and less than 1.0 in others. For
gle antenna, colonial algae have an antenna ar- brevity,wesaythatthedirectionoftheantenna
580 FOSTER AND SMYTH MICROBIOL. REV.
isthedirectionofmaximum directivity. Wefre- particle and a magnetic component that exerts
quentlyuse theworddirectivity simply torefer a force proportional to the velocity of the
tothedirectionalpropertiesoftheantenna.The charged particle. However, when light passes
antenna ultrastructures we discuss are speciali- through matter, the only effects we need con-
zationsthatachieveapatternofdirectivitysuit- sider are the electric forces on electrons. Light
ableforphototaxis. exertsaperiodic force on the electrons ofevery
Allalgalantennasaredesignedtoseeprimar- molecule in its path, setting the electrons into
ily in the forward direction. For the most part, oscillation at a frequency equal to that of the
their field of view is limited to light beams light. Ifthisfrequency is the natural oscillating
coming from the front of the antenna. In this frequencyoftheelectrons,aphotonoflightmay
respect they are analogous to man-made aper- be absorbed. Otherwise, the oscillating charges
tureantennas, anditwillbeusefulindiscussing cause light to be emitted in all directions from
algal antennas to define their effective aperture themolecule. Thisshowsthenetvelocityofthe
asthesizeofthedetectornormaltothedirection lightwavesuchthatthevelocityofpropagation
of the antenna. It is also useful to define a vis
measure ofthe angular acceptance oflight, the
half-beamwidth.Thisistheanglebetweenhalf- v= c/n (2)
intensity points in the main lobe ofdirectivity wherecisthevelocityoflightinavacuum, and
(angle a in Fig. 9a). All algal antennas are also n isdefined asthe refractive indexofthe mate-
analogoustoso-calledscreen-reflectorantennas, rial.
which have an absorbing and reflecting screen Inmoleculesinwhichtheelectronshavebeen
behind the sensing element. To evaluate the raised to a higher energy level by absorption of
effectivenessofthisscreen, we have introduced a photon, the light intensity diminishes during
a measure ofthe front-to-back contrast, some- passage through the material. The two effects,
timescalledthemodulationorcontrastfunction, reduction in velocity and attenuation of inten-
definedas sity,aredescribedquantitativelybythecomplex
refractive index
m = (Ifront -Iback)/(Ifront +Iback) (1)
Valuesofmrangefrom0fornocontrastto 1for n = n - ki (3)
completecontrast. Inourcase,Ifrontisthedirec- wheretherealandimaginaryindicesnandkare
tivity in the antenna direction, and Iback is the bothrealnumbers and i = ~,Pi.
directivity in the opposite direction. The con- Atthe interface between two media ofdiffer-
trastfunctionhasbeenausefulmeasureinsen- ent composition light is both refracted and re-
sorysystemsand elsewhere (37, 132). flected. The amountsare afunction ofn, k, and
Wehavesaidthatalgalantennasarethesame theangleofincidence. Thisisimportantinpho-
orderofsize asthewavelength ofthelight they totaxis, andmuch ofalgalultrastructure can be
receive.Asiswellknown,theapproximations of understood asingenious manipulations ofinter-
geometric optics are strictly valid only for ob- facestoachieve desirablelight distributions.
jectsthatarelargerelativetothewavelengthof Onefallacythathascausedunnecessary con-
light, but this should not discourage the reader fusion should be disposed of at the outset. It
from thinking about algal antennas in terms of mightbearguedthatachievementofdirectivity
more familiar optical devices. Even at sizes be- inalightantennaisatrivialtask.Theargument
low about five wavelengths across, for which is as follows. Imagine a thin flat membrane of
geometric optics is no longer quantitatively ac- area A cm2 containing pigment molecules ori-
curate,itgivesaqualitativepicturethatisuseful ented at random. Iflight ofintensityIphotons
andnotgrosslymisleading (121). cm-2 s-1 is shone on the membrane at normal
With this general picture of an antenna in incidence, IA photons will pass through the
mind,wepresentthedifferentopticalprinciples membranepersecond.Ifthemembraneistilted,
that apply to understanding algal antennas be- thequantumfluxbecomesIA*cosGphotonsper
fore we introduce the complicated structures second,where6istheangleofincidence.Tilting
themselves. the membrane reduces the number ofphotons
passing through it. From this, one might con-
OpticalPrinciples
clude that the number ofphotons absorbed by
All the optical effects we discuss depend on the pigment molecules will also change. This is
the interaction of light with matter. Popular false. The absorption does not change, because
accountsaregivenbyWeisskopf(246) andsome the probability that a molecule will absorb a
optical texts (56, 108, 130). Light is an electro- photon depends only on the intensity of light
magnetic wave, meaning that it has an electric striking each molecule and the average absorp-
component that exerts a force on any charged tion cross section of each molecule. Neither
VOL. 44, 1980 ALGAL PHOTOTAXIS 581
quantity changes with angle of incidence pro- number, we have a = 103 lnlO.E/Na = 3.82 x
vided that the number of photons absorbed is 10-21E.
small, in other words, that the pigment mole- IncalculationsinvolvingMaxwell'sequations,
cules do notscreen each other. Directivity can- whichdescribeelectromagneticwaves,thecom-
notbeachievedmerelybyputtingpigmentmol- plex refractive index (equation 3) is more con-
eculesinalayer.Theopticalprinciplesdiscussed venient. Fork, the complex term describingab-
below are necessary. sorption, wehave
Absorption. Mostphototactic algae are also
k = aX/4T (7)
photosyntheticandhaveahighconcentrationof
photosyntheticpigments.Inaddition,mosthave whereAisthewavelengthoflightinvacuum (in
a deeply pigmented organelle, the eyespot. Ab- centimeters).
sorption by the eyespot pigments is an impor- Scattering. All cells are optically inhomoge-
tant, but by no means the only, contribution of neous and therefore scatter light. Scattering is
the eyespotto antenna directivity. Forapurely certaintobeamajorfactoraffectingdirectivity,
absorbing screen, Iback = Ifront x T, where T is though the effect of scattering has often been
the fraction transmitted, so that the contrast neglected in discussions ofphototaxis. Forscat-
function teringwehavethe contrastfunction
m= (1 - T)/(1 + T) (4) m= (1+S- T)/(1+S+ T) (8)
Absorption oflight by a pigmented structure is where S is the fraction back-scattered, and Tis
always accompanied by a certain amount of the fraction transmitted. Scattering may also
reflection. decrease local contrast (as it does in Phyco-
Absorption canbe described inseveral differ- myces) by lighting otherwise dark areas ofthe
entways,themostusefuldescription depending cell.
onthe application. Wesummarize fourdescrip- Refraction. Itisnotknowninmostinstances
tions thatwewilluse later. whether refraction is significantly involved in
Lambert's law states that equal paths in the producing directivity. Cells and colonies ofcells
same absorbing medium absorb equal fractions are large enough to act as lenses. It is doubtful
ofthelightthatentersthem.Fromthisitfollows that the difference in refractive index between
that the cells and the surrounding water is great
enough to cause appreciable focusing at the far
T= eaex (5)
sideofthecell. Aroundtheequatorofthecellor
where a is the absorption or attenuation coeffi- colony a ring ofdecreased intensity is produced
cient (per centimeter), and x is the distance that might have a significant effect. The ocelli
through anabsorbinglayer (in centimeters). ofthe large Warnowiacean dinoflagellates (96),
For a solution of an absorbing substance, shown in Fig. lld, have a lens that does refract
Beer'slawstatesthatabsorptionisdirectlypro- thelightsufficiently tomaketheocellusadirec-
portional to the concentration. To the extent tional receptor (84).
thatthis istrue we have, astypically written, Reflection andinterference. The reflectiv-
ity or reflectance (ratio of reflected energy to
T= 10-ECX (6)
incident energy) whenlightpassesfromatrans-
where E is the molar extinction coefficient (in parent mediumto anabsorbingmedium atnor-
liters per mole per centimeter), and c is the mal incidence is
concentration of the absorbing compound (in R = [(n -n2)2 + k22]/[(n1 + n2)2 + k22] (9)
molesper liter). Itfollowsthata = lnlO.Ec.
Forourpurposes, the mostusefulmeasure of where ni is the refractive index ofthe transpar-
absorptionisusuallytheabsorptioncrosssection entmedium, andn2andk2 arethecoefficientsof
a (square centimeters). amaybeassumedto be the complexrefractive index (equation 3) ofthe
the area associated with each absorbing mole- absorbingmedium. Thecorresponding equation
cule,suchthatifaphotonhitstheareaitwillbe for light passing from an absorbing to a trans-
absorbed, otherwise itwillpassthrough. Stated parent medium is
differently,aistheprobabilitythatonepigment
molecule in athinlayerwith area 1 cm2 will be R = [(n,2 + k22- nln2)2 +(n2ki)j/ (10)
excitedbyonephotonpassingthroughthelayer [(ni2 + k12 + nln2)2 + (n2kA)1
(for example, see Clayton [40] or Blanchard et
al. [21]). The absorption coefficient is a = Nra, Forreflectionwithoutinterference, thecontrast
where Nt, (per cubic centimeter) is the number functionis
ofpigment molecules perunit volume. Because
N} = Nac/103 cm-3, where Na is Avogadro's m = (1 + R-T)/(1 + R + T) (11)