BACTERIOLOGICALREVIEWS, Mar. 1973, p. 32-101 Vol. 37,No. 1 Copyright0 1973 American Society forMicrobiology PrintedinU.SA. Physiology and Cytological Chemistry of Blue-Green Algae C. PETER WOLK MSU/AECPlantResearch Laboratory, Michigan State University, EastLansing, Michigan48823 INTRODUCTION ..................... 33 .............................. CYTOLOGY AND CYTOLOGICAL CHEMISTRY ......... 33 .................... Outer Layers ............... 33 .................................... D Sidewalls ............... 33 ..................................... o End walls; division ................................................... 34 w Mucilage/sheath ....................................................36 n Membranes ............. ...................................... 36 lo Plasmalemma ................... 36 a ................................ Photosynthetic lamellae (thylakoids) ................. 37 d ....................... e Pigments, lipids, andelementsofelectrontransportchains.38 d Gasvacuoles(pseudovacuoles) ...............................................41 f Other Intracellular Components ...............................................42 ro DNA ................................................... 42 m RibosomesandRNA.................................................... 43 h Glycogengranules(a-granules) .............................................. 43 t t Cyanophycingranules(structuredgranules) ........... 44 p ...................... : PPohloys-p,h-ahtyedrgorxaynbuulteysrat(evolgurtainnu,lemse.t.a.c.h.r.o..m.a.t.i.n.)..............................................................4455 //m Polyhedral bodies 45 m .................................................... Lipid deposits ................... 46 b ................................ r Phycobilisomes and biliprotein pigments .......... 46 . ........................ a Otherinclusions.................................................... 49 s OverallChemicalComposition ................................................. 49 m PHYSIOLOGY ............... .................................... 50 .o PhotosyntheticLightReactions ................................................ 50 r g Intactcells ............... .................................... 50 / Cell-free systems .................................................... 51 on CarbonMetabolism....................................................53 Pathways ofCO2 fixation and carbohydrate breakdown ....... 53 A ............. p Acetate metabolism....................................................54 r Respiration ............. ......................................55 il Intact cells ............... 55 1 .................................... , NADH andNADPH(etc.) oxidation .................. ....................... 55 2 Interactions with photosynthesis............................................. 55 0 Nitrogen Metabolism.................................................... 56 19 Nitrogen (N2) fixation ................................................... 57 b Reduction ofnitrate and nitrite ............................................. 57 y Incorporation of ammonia .................................................. 57 g Aminoacidmetabolism ................................................... 58 u Nucleicacidmetabolismandproteinsynthesis .......... 58 e ..................... s Sulfur Metabolism....................................................58 t Genetics ................................................... 58 Cyanophage .............. 59 ..................................... Movement ............ 60 ....................................... Mechanism ............... 60 .................................... Movement in the dark; effects ofnonphotic parameters ....... 61 ............. Effects of light on movement: changed speed of movement ...... 61 .......... Orientation relativetothedirectionoflight ........... 61 ...................... Response to agradient ofintensity ................... 62 ....................... Culture .62 Inorganic nutrition..62 32 VOL.37,1973 BLUE-GREEN ALGAE 33 Organic nutrition. 64 Light and temperature. Synchronous growth.66 Means of obtaining pure cultures.66 Growth of single cells .66 Substances ted; Toxins.66 Regulation and Development..67 Biochemical regulation..67 Chromatic variation..67 Development ofNostoc muscorum,..68 Spores (akinetes)...68 Intercellular interactions; control of patternization .69 Heterocysts.70 EVOLUTION.73 Blue-green Algae and OtherProkaryotes.73 OriginofChloroplasts.76 D LITERATURECITED....... .77 o w n lo INTRODUCTION CYTOLOGY AND CYTOLOGICAL a d CHEMISTRY e d This review grew out ofan interest in devel- Outer Layers fr opmental phenomena in certain filamentous o blue-green algae (823-825). In these algae, Side walls. The structural features of the m vegetative cells can differentiate into hetero- cells of blue-green algae-or Cyanophyta- h t cysts, andsometimes alsointospores.Differen. which can be observed with the light micro- tp tiation involves majorchanges incertainofthe scope include a central region ("centroplasm, :/ / cytochemical and physiological characteristics nucleoplasm") which is rich in nucleic acid, m of the vegetative cells, whereas other such and which interdigitates with a peripheral m characteristicschangelittleifatall.Afirststep region ("chromatoplasm") containing thepho- b r toward understanding the transition from the tosynthetically active pigments; various inclu- . a metabolic state ofavegetative cell to that ofa sions; and circumferential layers including a s m heterocyst or spore is to learn the characteris- plasmalemma, a pellicular wall, and often, a . tics ofthose end points. layer of mucilaginous material (250). Figure 1 o r The positions of the heterocysts and spores is asurvey electron micrograph ofablue-green g / along the algal filaments form well-defined algal cell, showing with greater resolution the o patterns. These developmental patterns, at features cited above. The following sections n least in Anabaena cylindrica, are dependent will discuss the fine structure and chemical A upon interactions between cells, as well as composition of the component parts of these p r intracellular changes. Study of development cells, beginning with the cell wall. il 1 may therefore be expected to be assisted by Betweentheplasmalemma andthe extracel- , consideration ofother phenomena, e.g., move- lular mucilage is a wall layer, termed the 2 0 ment and nitrogen fixation, which involve co- "inner investment" (250), which has been re- 1 ordinatedactivitiesof,andwithin, cellsofalgal solved by the electron microscope into four 9 filaments. layers, LI through L1, (438; Fig. 1). This b y Thus, a survey of information pertinent to terminology appears now to be generally ac- g study of differentiation and pattern formation cepted (e.g., 12, 335, 484). LI,, is discussed u in blue-green algae expanded to encompass below. LayersLIandLII,areelectrontranspar- e s muchoftheirphysiology andcytologicalchem- ent, and vary from about 3 nm each in thick- t istry. This review is perforce a primer because ness (429) to about 10 nm (438). L1 may ofthe restrictions ofcontemporary knowledge. possibly be an artifact of preparation (12; but It is written with the hope that it will help to cf. 107). Aparallel arrayoffibrils, 6to9nm in "prime" the maturation of developmental diameter, is present in the region of, and studies ofthese algae. apparently constitutes, layer L;j, of Oscilla- In recent years, Lang (484) has reviewed the toriaprinceps(335).ThethicknessofL1 ranges cytology, and Holm-Hansen (390) the physi- from 10 nm or less in Oscillatoria rubescens ology and ecology of blue-green algae. Other (438) andAnacystisnidulans(516)toabout200 reviewswillbecitedwheretheyarepertinent. nm in 0.princeps (335). Thislayerthickens at 34 WOLK BACTERIOL REV. least fivefold during akinete differentiation in dark pattern, typical of "unit membranes" Cylindrospermum sp. (427). following permanganate fixation (438), or the Rows of pores are frequently visible on both image ofonly asingle darkline (200). Undula- sides of the loci of ingrowth of end walls, in tion of this membrane (107, 200) may be an filamentous blue-green algae.Theseporesare7 artifact, since its appearance is smooth follow- to 20 nm in diameter (179, 543), or may be ingfreeze-etching (438; cf. also 657). Alayerof elongated at the outside (616), and may taper similar location and appearance in gram-nega- toward the inside (616) or toward the outside tive bacteria consists of lipopolysaccharide (335). InMicrocoleus vaginata,hormogonepro- (606), and it has beenfoundthat2to3%ofthe duction involves tearing ofthe wall alongjust dry weight of Anacystis nidulans is a wall- such a circumferentially positioned setofpores associated lipopolysaccharide of which about (482). It has been suggested, but with little 60% is carbohydrate, principally mannose but supporting evidence, that these pores are in- with glucosamine, 2-amino-2-deoxyheptose, 2- volved in mucilage secretion (179, 543). How- keto-3-deoxyoctonate, and other sugars also D ever, no such pores were seen in a study ofthe present (800). ow ultrastructure of members of the Based on the content ofmannose and amino n Stigonemataceae (107). It was suggested that sugars in cells and walls, Dunn et al. (191) lo the mucilage ofthesealgaemaybeextruded as estimated that 10 to 20% of the dry weight of a d "blebs" ofmaterial. vegetativecellsofA. cylindricaiswall material e Blue-green algae can be lysed by growing (exclusive of mucilage). Walls of blue-green d them in penicillin (e.g., 237, 241, 254, 470), algae contain a variety of amino acids and fr o which interferes with deposition ofpeptidogly- thus, presumably, protein (388, 643). m can into bacterial walls. Moreover, lysozyme, End walls; division. The end walls of uni- h which breaks down this material in bacterial cellular blue-green algae do not differ in ultra- t t walls, can cause lysis ofblue-green algae (254), structure from thesidewalls(e.g., 12, 200, 420). p : a fact which has permitted isolation of proto- In filamentous species, a peptidoglycan-con- // m plasts (61, 161, 779) or spheroplasts (187, 276, taining (241) L11 layer is present, sandwiched m 331, 633) of these algae. Since simultaneous between LI layers which are continuous from b treatment with ethylenediaminetetraacetate theside walls ofthe two adjacentcells (438; cf. r . (EDTA) (187) or prior lyophilization (297) also332, 333, 506).Theresultisathree-layered a s sometimes greatly increases the effect oflyso- end wall (254, 332, 333, 438, 572). In m zyme, it appears that the sensitive layeris not Stigonema, the Li, layer may subsequently be . o directly at the cell surface. Although initial dividedpartofthewayinfromtheperipheryof r g resultssuggestedthatL,wasthelysozyme-sen- the end wall by an ingrowth oflayers L,,1 and / sitive layer of the cell wall (241), it has now LI, (107). o n been clearly demonstrated that the layer de- Pores (10-20 nm) were observed in chromic A stroyed by lysozyme is LI, (429; cf. also 516). acid-treated end walls of Oscillatoria sancta p pepAtftiedrogiltycwaans(f6o70u,nd730t)h,atintchleudcionngstdiitaumeinntospio-f acrnodcolienuse,ndanwdalOlssciolflatCoyrliiandlrimoosspaer(m1u7m9,, 1M8i0-, ril 1 melic acid (831; and cf. 395), are present in 543). Hagedorn (333) found 35 to 40 pores per , 2 blue-green algae, Frank et al. (240, 241) and ,gm2. Intercellular "plasmodesmata" 17 nm in 0 later workers analyzed the composition of the width were observed in thin sections of Sym- 1 9 peptidoglycan-containing layer of the wall plocamuscorumbyPankratzandBowen(616). b (Table 1). There is little divergence from the It is unknown ifthese structures play any role y ratio of constituents expected ifthe backbone in intercellular communication. g of the peptidoglycan is, as in most bacteria Theendwallsareformedbyirislikeingrowth u e (606), a repeating sequence of N-acetyl mu- of the side walls (332), beginning with the s ramic acid and N-acetyl glucosamine. plasmalemma (616). Ingrowth of the Lj1 layer t Carbohydrates detected in isolated walls of may beeither as acontinuation oftheL,,layer blue-green algae are listed in Table 2. of the side walls (333) or from a ring-formed Some authors have pointed outtheexistence deposit(438),suchasissometimesfoundatthe ofamembrane-like layerL1\, approximately 75 junction ofcompleted end wallswithsidewalls to 80 A thick, external to the peptidoglycan (252, 572) in filamentous forms. Division in layer ("outer membrane," reference 200; Agmenellum quadruplicatum strain BG-1 "Scheidenlamella," reference 438; see also takes place by simultaneous invagination ofall 654). This layer is, however, not always seen wall layers (420), and in Anacystis nidulans, (654). The layer may have either a dark-light- another unicellular blue-green alga, by forma- VOL.37,1973 BLUE-GREEN ALGAE 35 PI LI LII LIU LIVM 1.II1 J,.,r.,.11 F- A !,;. 4 A i '..'ite-. .16. D o w n lo a d e d f r o m h t t p : / / m m b r . a s m . o r g / o n A p r il 1 , 2 0 1 9 b y g u e s t FIG. 1. A survey electron micrograph ofa cell ofAnabaena variabilis showing mucilage (M); wall layers (L-Lwv); plasmalemma (P1); thylakoids (T); DNA-containing regions (N), with fine fibrils (F) visible in certainareas (seeinset); cyanophycingranules (CG); apolyhedral body (PB); andotherinclusions,possibly lipid innature (L). Courtesy ofL. V. Leak (500); withpermission. x48,000.Inset, x272,000. 36 WOLK BACTERIOL REV. TABLE 1. Composition ofpeptidoglycanfrom various blue-green algae Peptido- Molarratioa glycan! Organism IsWWoallaallt.led Mura.mic Gluc.os- Dpipiiammmeeillniioc- Gluta.m~ic AAllaanniinne AsparticRRefeerrnencce (%wt) acid amine acid acid acid Phormidiumuncinatum 52 0.63 1.0 1.0 0.94 2.0 0.94 241 P.foveolarum 32 0.8 1.1 1.0 1.1 1.85 388 Anacystisnidulans 28 1.47 1.34 1.0 1.52 2.4 187 A.nidulans 22 0.9 1.9 1.0 1.1 2.0 388 Tolypothrix tenuis 25-30 0.8 1.1 1.0 1.1 1.9 388 aDiaminopimelic acid = 1. D o w TABLE 2. Sugars in wallpolysaccharides from various blue-green algae n lo Sugars a d Organism MMaann-- Glucose GGaallaacc-- Xylose Ribose Arabi- Rham- Fucose Gtaolsaac-- Reference ed nose tose nnoossee nnoossee mine kknoown f r o Phormidium m foveolarum + + 388 h Tolypothrix tt tenuis + + + + 388 p: Anacystis // m nidulans +a + + + + + + + 388 m A.nidulans +++ + + 186 Anabaena b r cylindrica .a (vegetative s m cells)b 50% 35% 5% 8% 2 1% 192 . o aA highly purified peptidoglycan fraction was still rich in mannose. rg hCarbohydrate accounted for ca. 18% ofdetergent-washed walls. / o n tion of a three-layered end wall (see above), kali.) Itisnotpresentlyunderstood, inchemical Ap wahnidcLhIi,sltahyeenrstr(a1n2)s.ected byingrowthoftheLI1I atlegrames,arwehdyiffcleuretnatin(19m2u),cilwahgeerseasofotbhleure-mgurceie-n ril 1 When a cell ofPleurocapsa fuliginosa forms lages-referred to as sheath (729)-are much , endospores, many small groupings, each of denser. 20 which contains thylakoid fragments and a Substances, presumably mucilaginous poly- 1 nuclear region, become distinguishable within saccharide, produced by Anabaenaflos-aquae, 9 the cell. These groupings arethen separated by and capable of reducing friction (drag) in by ingrowth ofthe cell wall (49). turbulent flow, have a molecular weight esti- g Mucilage/sheath. A layer of mucilage sur- mated to be about 100,000 (417). u e rounds the cells or filaments of many blue- s green algae. In shadow micrographs (248, 543, Membranes t 685, 708) and in thin sections (481, 501, 775), Plasmalemma. Surrounding the protoplast mucilage has a fibrillar appearance, but the and internal to the cell wall is a plasma presence offibrils 30Awide, andwider, maybe membrane or plasmalemma, demonstrable at due to binding together ofindividual fibrils by the light microscope level, albeit with diffi- the act of drying (438). Chemical studies of culty, by plasmolysis (137, 250). This boundary presumptive mucilage aredescribedinTable3. (Fig. 1) is variously estimated as 70A (616) up (It isdifficulttoascertaintheoriginofallofthe to 100 to 150 A (200) thick. The plasmalemma material extracted with hot water or hot al- is apparently not the sole permeability barrier VOL.37,1973 BLUE-GREEN ALGAE 37 TABLE 3. Compositionofputativelyextracellularmucilaginouspolysaccharidesfromvariousblue-greenalgae Monomers Aiulgria 100 MHo2deof + Refer- extraction aC.0 aD 0 ~ eencc 0 a ') 0-~-~~'-'~.a ~CZ0 Rivularia 100C,H20 + + +?623 Calothrix 100C,H20 +? + +? 623 Phormidium tenue 100C,H20 + +? +? + + 624 Nostoc 100C,H20 + 20- 25% 10% 30% + 415 D Anabaenaflos-aquae Ethanolpre- 67% 30% 30% 2.3% 0.8% 553 o cipitation w Anabaena cylindrica 100C,4% n (Foggstrain) NaOH 31% 0 6% 25% 6% 6% 25% 68 lo Anabaena cylindrica Centrifugation, a d (Wolk strain) ethanol wash 47%a 25% 6% 21% trace 0 0 O 192 e d aThese figures refertothe polysaccharides; 5 1% amino compounds andca. 7% ashwere alsofound. fr o m in algal cells, since protoplasts ofA. nidulans On occasion, reticulate configurations ofla- h were found to take up nucleic acid precursors mellae are seen (487, 541; cf. also 485). Al- tt more rapidly than do intact cells (633). In- though most often seen closely appressed, the p: stances of apparent fusion of photosynthetic thylakoid discs can swell greatly (225, 654, //m lamellae to the plasmalemma have been re- 806), a condition apparently correlating with m ported (11, 616, 681) but may have arisen as growthathighlight intensity (225, 806). Inthis b artifacts of fixation and sectioning. However, case, the "vacuolar space" within the disc is r . following degeneration of lamellae in 0. ru- aqueous (438). The plane of the membranes as bescens, renewed formation of lamellae origi- canbe parallel ororthogonal to thecellsurface m nates in invaginations from the plasmalemma. (572). Although lamellae usually appear to be .o These in turn develop secondary invaginations both numerous and discrete within a cell, rg whichgrow andthendisperse as smallvesicles. instances are known in which only a single / The vesicles enlarge intophotosynthetic lamel- peripheral lamellaisfound (43), andinwhicha on lae (438). Structures of appearance similar to single thylakoid spirals around the centro- A the invaginations mentioned havebeen likened plasm (337; cf. also 485). p toPmheostoossoymnethset(2i0c4,l6a2m9e)l.lae (thylakoids). Al- deJnocset t(h4a3t8)thheasouptreersenftaecde porfeltihmeinlaarmyelelvaie- ril 1 though detectable with the light microscope consists of50 to 70 A subunits; that 100 to 200 , 2 (292), the photosynthetic lamellae ofthe blue- Aparticles, relatively thickly packed, arepres- 0 green algae were seen first with the electron ent on the inner face of the membrane; and 19 microscope, initially in shadowed preparations that these are separated by an "unstructured" b (108) and later in thin sections (572, 573). membrane. In contrast, Fuhs (256) has main- y The lamellae are closed discs (332, 438, 654) tained that the membranes are comprised of gu termed thylakoids (541; Fig. 1). The appear- spherical subunits. It would appear that the e ance of the lamellae, when adjacent mem- detailed substructure ofthe membranesmerits st branes are appressed, varies with the fixative much further investigation. employed. Following permanganate fixation, All, or almost all, ofthe cellularchlorophyll, the appearance ofa double membrane, asseen and much or all of the carotenoids, are local- in section, is ofthree parallel linesofthickness ized in the lamellae (11, 693), which may be 20, 40, and20A,betweenwhicharespacesof30 isolated by differential centrifugation (693, to35A (506, 616), whereasfollowingosmicacid 703). Schmitz (677) has presented a detailed fixation such a length appears as two parallel chemical analysis of thylakoids from Oscilla- lines, each 35 5 A (506) to 65 5 A (616) toria chalybea, including an amino acid analy- thick, separated by a space of about 50 A (cf. sis of their protein. Since chlorophyll ac- also 654). counted for 2.2% of the dry weight of intact 38 WOLK BACTERIoL REV. (a) H CH3- CH3 D o w (b) (c) n lo a d Hi e CH3 -CH3 CH3 -CH3 d fro IOOCCH2CH2- :CH-CH3 m HOWD I-CH3 h t t p HOOCCHZCH2 : HOOCCH2CH2 // m m FIG. 2. Tetrapyrrole pigments of blue-green algae. (a) Chlorophyll a, (b) phycocyanobilin, and (c) phyco- b r erythrobilin. . a s m . o cells and for 8.5% ofthe dry weight ofisolated transport chains. The only chlorophyll found r thylakoids, the thylakoids accounted forabout in blue-green algae is chlorophyll a (e.g., 544, g/ 26%ofthedryweightoftheintactcells(cf.also 556, 691, 692; Fig. 2). Ithasbeenconfirmedthat o n 754). the chlorophyll from Phormidium luridum con- A Thornber (761) solubilized a chlorophyll a- tains phytol (728). In vivo, theabsorption peak p pwriottheisnodcioummpledxodefcryolm sPuhlfoartem.idTihuem cloumrpildeuxm tishasnhifttheed taobsaorwpatvieolnenpgetahkabinoutme1t3hannmolhi(g2h0e9,r ril 1 contains 70% of the chlorophyll a of the orga- 703). Representative values of chlorophyll as , nism, is homogeneous upon gel electrophoresis percent of algal (dry weight) are 2.2 (677) and 20 and ultrafiltration, and contains four subunits 0.2to1.0(741). 1 (mol wt ca. 35,000; complex mol wt ca. Tabulations of the quantities of individual 9 160,000). Per subunit there are five chloro- carotenoids found in a large variety of blue- by phyll molecules plus traces of 13-carotene and green algae are presented in the papers of g 4-keto-fl-carotene. It appears that one-fifth of Stransky and Hager (741) and Hertzberg et al. u e the complexes contain P700; a quinone is also (375). Suffice it here tosaythat 13-carotene (i), s present (175). Ogawa et al. (597), having dis- echinenone (ii), myxoxanthophyll (xi), and t rupted lamellae fromAnabaena variabilis with sometimes zeaxanthin (viii) are most fre- Triton X-100, found a dense particle contain- quently the major carotenoids present, al- ing P700 and a less dense particle with myxo- though in exceptional cases canthaxanthin xanthophyll. Whether the latter particle is re- (iii), caloxanthin (ix), nostoxanthin (x), oscil- latedtophotosystemII (597) isunclear, butthe laxanthin (xii), and aglycoside (xiii) havebeen presence ofP700 in complexes prepared bythe found to account for 10% or more of the total two groups ofworkers indicates the association carotenoid present. The structures of the ca- ofthose complexes with photosystem I. rotenoids are as follows. Pigments, lipids, and elements ofelectron (i) 13-Carotene (Fig. 3i) is the only carotene VOL.37,1973 BLUE-GREEN ALGAE 39 (x) Nostoxanthin (3,3'-dihydroxy-5,5'-dihy- () dro-7,7'-didehydro-f,-carotene) (see ix). (xi) Myxoxanthophyll (1',2'-dihydro-3',4'- , -carotene didehydro-3,1'-dihydroxy-y-carotene= "myxol," with rhamnose or, as a minor compound, HOX hexose in glycosidic linkage at the 2' position; Fig. 3xi). Myxoxanthophyll, a major xantho- phyll in most blue-green algae examined, has caloxonthin (but seetext) not been detected in other members of this group (375). Its structure was established by Hertzberg and Jensen (372). (xi) N N rhamnose (xii) Oscillaxanthin (1,1'-dihydroxy-2,2'-di- HO0 OH - L - rhamnosyl - 1,2,1',2' - tetrahydro-3,4, D myxoxanthophyll (myxol-2'-rhamnoside) 3,4'-tetradehydrolycopin 2,2'-dirhamnosyl - ow "oscillol"; Fig. 3 xii; reference 373). n OHN (xiii) Myxol-2'-O-methyl-methylpentoside, lo rhamnose0 0rhamnose as also xiv and xv, was isolated from 0. limosa ad (Xii) O and assigned this structure by Francis et al. e oscillaxanthin(oscillol-2,2'-dirhamnoside) (238). d f (xiv) Oscillol-2,2'-di-(O-methyl-methylpen- ro toside) (see xiii). m (xvi) o (xv) 4-Keto-myxol-2T-methylpentoside (see h xiii). tt mutatochrome p (xvi) Mutatochrome (flavacin; Fig. 3 xvi; : FIG. 3. The basic structures ofcarotenoids ofblue- // greenalgae. reference 371). m (xvii) Aphanizophyll has spectral properties m very similar to those of myxoxanthophyll. b r which has been foundin blue-green algae (e.g., Nonetheless, the two pigments are not identi- .a 120, 320, 363, 447, 479, 763). cal (370). Aphanizophyll may be 4-hydrox- s m (ii) Echinenone (myxoxanthin; aphanin; 4- ymyxoxanthophyll (374). . keto-f3-carotene), "myxoxanthin" (363), and (xviii) Unknown carotenoids have been o r "aphanin" (763, 764) have been shown to be found in trace amounts in several blue-green g / identical to 4-keto-f3-carotene ("echinenone") algae(361a, 741). o (319, 321, 370). The principal water-soluble pigments of n (iii) Canthaxanthin (aphanacin; 4,4'-diketo- blue-green algae are discussed below, under A #-carotene; reference 370). "phycobilisomes and biliprotein pigments." pr (iv) 4-Keto-4'-hydroxy-3-carotene (741). Mono- and digalactosyl diglyceride, phos- il (v) 4-Keto-3'-hydroxy-f3-carotene (369). phatidyl glycerol, and sulfoquinovosyl diglyc- 1, (vi) Cryptoxanthin (3-hydroxy-0-carotene; eride (Fig. 4), the four principal fatty acid-con- 2 0 reference 741). taining lipids found in chloroplasts, are also 1 (vii) Isocryptoxanthin (4-hydroxy-fl- present in blue-green algae. Lecithin, phos- 9 carotene; reference 361a). phatidyl ethanolamine, and phosphatidyl ino- b y (viii) Zeaxanthin (3,3'-dihydroxy-fl-caro- sitol have not been found in these algae (567). g tene). Originally identified as lutein ("leaf Tabulations ofthe abundance ofdifferentfatty u xanthophyll"), an intensely orange compound acids, principally C14, C1s, and C18, in a e s in extracts of 0. rubescens was subsequently variety of blue-green algae may be found in a t shown to be zeaxanthin (369, 447), and the number ofpublications (399, 452, 567, 620). presence of zeaxanthin and absence of lutein The absence of a-linolenic acid (18:3 [3, 6, have been confirmed forotherblue-green algae 9]) from A. nidulans and certain other blue- (320, 741). green algae (398, 452, 453, 567, 620, 729) shows (ix) Caloxanthin, as also (x), was first ob- that, although a-linolenic acid is common to served inA. nidulans (379, 741). The structure eukaryotic photosynthetic plants, it cannot be 3,3' - dihydroxy - 5 - hydro - 7 - dehydro - fl - a requirement for the photosynthetic produc- carotene (Fig. 3ix) was assigned by Stransky tion of oxygen. However, a-linolenic and (or) and Hager (741). The validity ofthis structure, -y-linolenic acid (18:3 [6, 9, 12]) is present in however, has been questioned (375). other blue-green algae (452, 453, 512, 567, 568, 40 WOLK BACTERIOL REV. 620, 729). The absenceofa-linolenic acidisnot 0 restricted to unicellular blue-green algae: Ha- CH3 1 palosiphon laminosus, which has a branched, Plastoqulnone A CH3 H filamentous habit, also lacks fatty acids with I0I 9 three double bonds (399, 568). Branched-chain CH3 a R CH3 fatty acids are almost completely absent from HO blue-green algae (620). Capric acid (10:0), Tocopherols R z H elsewhere rare, cancomprise asmuch as50% of CH3 the fatty acids of Trichodesmium (620). (Dis- CH3 tinctive lipids of heterocysts are discussed below.) a -Tocopherol- CH quinone 3rH I C15-C19hydrocarbons arefoundinblue-green CH3 algae (818), with n-C17 often predominating 0 D (295, 341), butwith C19constituting98to 100% - CH3 o of the hydrocarbons in three marine strains Phylloqulnon. N H w (818). Branched alkanes, principally 7- and 0 n lo 8-methyl heptadecane, have been detected in FIG. 5. Quinones and tocopherols of blue-green a extracts ofNostoc (342). algae (761a). Plastoquinones B and C aresimilar to d Sterols have not been definitively demon- plastoquinoneA, andhave theside chainhydroxyla- ed strated in blue-green algae (120, 511); where ted. In plastoquinone B, the hydroxyl group in the f side chain is acylated. r traces ofsterols were detected (557, 648, 727), o m no adequate proof was presented that eukar- yotic contaminants were wholly absent from h Both a- and ,B-tocopherol are present in A. t the cultures extracted. variabilis; neither is found in A. nidulans. Of tp Elements of electron transport chains and the two, only a-tocopherol is found in Nostoc :// otherpigmentsarediscussed herebecausethey m muscorum and Fremyella diplosiphon, and may be (but are not in each case necessarily) m from <0.2to 1.5,ugofa-tocopherolisfoundper structurally or functionally associated with b g (dryweight) inseveral otherblue-green algae r membranes. (115, 169, 366, 636). Incorporationoflabelfrom .a The quinones ofblue-green algae (see Fig. 5) s tyrosine and methionine into quinones and m include plastoquinones (PQ) A, also called PQ tocopherols of A. variabilis has been demon- . 9 (5, 100; see also 206, 510), B, and Cl6 (742); o strated (79). r the naphthoquinone, vitamin K1 (also called g phylloquinone), and (to date, found only inA. Cytochromes solubilized from blue-green o/ nidulans) a monohydroxy analogofvitamin K1 algae by sonic treatment are briefly summa- n (5, 115, 510, 742; see also 206); and membrane- rized in Table 4. These cytochromes are de- A scribed extensively in a series of papers by p bound a-tocopherolquinone (116).Atabulation r of the amounts of PQ A, vitamin K1, and Holton and Myers (400-402). A c-type cyto- il chrome was purified from Synechococcus, and 1 a-tocopherolquinone in five blue-green algae , hasbeenpresentedbyCarretal. (115),whodid its amino acid composition was determined 2 (162). 0 not detect ubiquinone in any ofthe algae. 1 In addition to cytochromes solubilized by 9 Lipid R sonic treatment, A. variabilis contains two b Phosphatidyl glycerol HOCH2CHOHCH20P(OH)20- cytochromes, absorbing at 557 and 562 nm, y g which remained tightly membrane-bound (596 u RICOOCH2 H2CSO5 and cf. 62, 744). Diphenylamine markedly in- e Sulphoquinovosyl diglycerIde s creases the cellular content of the 554- and t R2COOCH 562-nm cytochromes (596). OH Afterinitial work withA. nidulans andother R-CH2 Monogalactosyl diglyceride H1HCOOH blue-green algae (71, 400), ferredoxin was crys- tallized from Nostoc (549; cf. also 28). Exten- sively purified ferredoxin fromA. nidulans was HaCOH Digalactosyl diglyceride HO 0 characterized (835) asmolwtabout11,000with RlR2-fatty O-CHz two iron atoms and one labile sulfur atom per acids molecule (cf. also 212, 716, 744). Its amino acid composition has been determined (162, 835). FIG. 4. Lipids ofblue-green algae (566). Smillie (716) found that extracts ofA. nidu- VOL.37, 1973. BLUE-GREEN ALGAE 41 TABLE 4. Cytochromes solubilizedfrom blue-green algae Absorptionbands reduced(nm) Potential Comment Reference (EO V) No. a s Soret I i 553 521 416 0.30 c-Type from Tolypothrix 448;andsee162 tenuis 554 0.35 0.01 FromAnacystis nidulans 400;andsee596, 716,744 ii 549 -0.26± 0.02 c-Type; 400;andsee596, 716 Chlorophyll 220 (ii) = 1 Ferredoxin 0.86 D iii 552 (iii)/(ii) - 1:100 400;andsee596 o w n lo lans contain, in addition toferredoxin, aflavo- small peak in the action spectrum of 14CO0 a d protein-which hecalledphytoflavin-capable fixation at 719 nm. These phenomena are not e of efficiently mediating photosynthetic reduc- understood. d tion of oxidized nicotinamide adenine dinu- A pigment capable of photooxidizing uric fr o cleotide phosphate (NADP), despite the fact acid in vitro, and having an absorption max- m (81) thatitsredoxpotentialisgreaterthanthat imum at 730 nm in 50% ethanol, was isolated h of NADP. Phytoflavin lacks iron (716); its role from A. nidulans (33). This pigment is proba- t t may be to replace ferredoxin under conditions bly identical or closely related to a pigment p : of iron deficiency (81). Ferredoxin-NADP re- purified from Synechococcus cedrorum (286), // m ductase fromA. variabilis isalsoaflavoprotein and is probably responsible for mediating the m (744). An unidentified flavoprotein from photooxidation (most effectively at 750 nm) of b Synechococcus has been purified, and its uricacid, imidazole, etc. byA.nidulans(33; cf. r . aminoacidcomposition wasdetermined (162). also 326, 327). Whether a similar pigment is a s A copper-containing protein capable of act- involved in the slow, bleached growth of A. m ing as a redox reagent was purified from A. quadruplicatum on uric acid is unknown (40). . o variabilis by Lightbody and Krogmann (515) Gasvacuoles(pseudovacuoles). Theability r g andwas called "plastocyanin," thenamegiven of certain blue-green algae to float accounts / to similar proteins from photosynthetic tissues probably for their biological success, and cer- o n ofotherplants(seealso62). tainlyformuchoftheircontributiontoair-and A From 0.05 to 0.1% of the dry weight of A. water-pollution problems (233). Early observa- p nidulans is a tetrahydrobiopterin glucoside tions, recently reviewed (788; cf. also226, 250), r tentatively identified as having a dehydrated showed that, when suspensions of these algae il 1 side chain to which glucose is attached by an are subjected to a sudden pulse ofhydrostatic , 2 a-linkage (236, 530). The biochemical origin of pressure, the algae can no longer float. At the 0 theside chainhasbeenstudied (529). However, same time, certain reddish vacuoles of low 1 whether precisely this compound occurs and is refractive index disappear. The principal find- 9 the sole pteridine compound in A. nidulans is ing that led Klebahn (457, 458) to conclude by presently controversial (839). Pteridines were that the vacuoles are filled with agas wasthat g also found in members of the Nostocaceae the total volume ofalgal suspension decreased u e (236). upon disappearance of the vacuoles, and the s A heat-stable water-soluble factor, appar- change in algalbuoyantdensity wasconsistent t ently the same as one found in P. luridum and with substitution of liquid for gas in the vacu- capable of stimulating photosynthetic phos- ole volume. The interpretation that the con- phorylation of adenosine diphosphate (ADP), tents are gaseous was tested directly by Jost was isolated from spinach and found to be and Matile (441), whomadefreeze-etch micro- similar in spectral properties to an aromatic graphs of 0. rubescens frozen to -120 C, a pteridine (72; cf. also 277, 530, 531). temperature at which no water would subli- Dangeard (166, 167) observed a zone of mate. The vacuoles were empty, proving that growth, or perhaps accumulation, of they had contained a gas. Walsby (787) has Phormidium and Lyngbya at 720 to 730 nm, analyzed carefully the pressure relationships whereas Nultsch and Richter (588) found a involved ingas-vacuolerupture, andhasshown