Meissner M, Stark K, Cresnar B, et al.

Volvox germline-specific genes that are putative targets of RegA repression encode chloroplast proteins
CURR GENET 36 (6): 363-370 DEC 1999

Abstract:
In Volvox carteri, regA acts as a master gene to suppress all germ cell functions in somatic cells. Its product, RegA, has features of a transcriptional repressor. Here we report cDNA sequences representing 15 nuclear genes with properties expected of RegA targets: they are expressed strongly in germ cells and in regA(-), but not regA(+), somatic cells. Two of them encode polypeptides with no recognizable features, but ten (like three previously sequenced ones) encode chloroplast proteins of known function, and the remaining three encode putative chloroplast proteins of unknown function. This suggests that RegA blocks reproductive development in somatic cells by preventing chloroplast biogenesis, thereby making it impossible for the cells to grow enough to reproduce

 

Ender F, Hallmann A, Amon P, et al.

Response to the sexual pheromone and wounding in the green alga Volvox: Induction of an extracellular glycoprotein consisting almost exclusively of hydroxyproline
J BIOL CHEM 274 (49): 35023-35028 DEC 3 1999

Abstract:
The extracellular matrix (ECM) of Volvox is modified during development or in response to external stimuli, like the sex-inducing pheromone. It has recently been demonstrated that a number of genes triggered by the sex-inducing pheromone are also inducible by wounding. By differential screening of a cDNA library, a novel gene was identified that is transcribed in response to the pheromone. Its gene product was characterized as an ECM glycoprotein with a striking feature: it exhibits a hydroxyproline content of 68% and therefore is an extreme member of the family of hydroxyproline-rich glycoproteins (HRGPs), HRGPs are known as constituents of higher plant ECMs and seem to function as structural barriers in defense responses. The Volvox HRGP is also found to be inducible by wounding. This indicates that the wound response scenarios of higher plants and multicellular green algae may be evolutionary related.

 

Kirk DL

Evolution of multicellularity in the volvocine algae
CURR OPIN PLANT BIOL 2 (6): 496-501 DEC 1999

Abstract:
Recent studies reveal that relationships among the volvocine algae are more complex than was previously believed. Nevertheless, this group still appears to provide an unrivaled opportunity to analyze an evolutionary pathway leading from unicellularity (Chlamydomonas) to multicellularity with division of labor (volvox). Significant progress in this regard was made in the past year when two genes playing key roles in volvox cellular differentiation were cloned, and clues were uncovered regarding their mechanisms of action.

Coleman AW

Phylogenetic analysis of "Volvocacae" for comparative genetic studies
P NATL ACAD SCI USA 96 (24): 13892-13897 NOV 23 1999

Abstract:
Sequence analysis based on multiple isolates representing essentially all genera and species of the classic family Volvocaeae has clarified their phylogenetic relationships. Cloned internal transcribed spacer sequences (ITS-1. and ITS-2, flanking the 5.8S gene of the nuclear ribosomal gene cistrons) were aligned, guided by ITS transcript secondary structural features, and subjected to parsimony and neighbor joining distance analysis. Results confirm the notion of a single common ancestor, and Chlamydomonas reinharditii alone among all sequenced green unicells is most similar. interbreeding isolates were nearest neighbors on the evolutionary tree in all cases. Some taxa, at whatever level, prove to be clades by sequence comparisons, but others provide striking exceptions. The morphological species Pandorina morum, known to be widespread and diverse in mating pairs, was found to encompass all of the isolates of the four species of Volvulina. Platydorina appears to have originated early and not to fall within the genus Eudorina, with which it can sometimes be confused by morphology. The four species of Pleodorina appear variously associated with Eudorina examples. Although the species of Volvox are each clades, the genus Volvox is not The conclusions confirm and extend prior, more limited, studies on nuclear SSU and LSU rDNA genes and plastid-encoded rbcL and atpB. The phylogenetic tree suggests which classical taxonomic characters are most misleading and provides a framework for molecular studies of the cell cycle-related and other alterations that have engendered diversity in both vegetative and sexual colony patterns in this classical family.

 

Funke RP, Kovar JL, Logsdon JM, et al.

Nucleus-encoded, plastid-targeted acetolactate synthase genes in two closely related chlorophytes, Chlamydomonas reinhardtii and Volvox carteri: phylogenetic origins and recent insertion of introns
MOL GEN GENET 262 (1): 12-21 AUG 1999

Abstract:
Acetolactate synthase (ALS catalyzes the first committed step in the synthesis of branched-chain amino acids. In green plants and fungi, ALS is encoded by a nuclear gene whose product is targeted to plastids (in plants) or to mitochondria tin fungi). In red algae, the gene is plastid-encoded. We have determined the complete sequence of nucleus-encoded ALS genes from the green algae Chlamydomonas reinhardtii and Volvox carteri. Phylogenetic analyses of the ALS gene family indicate that the ALS genes of green algae and plants are closely related, sharing a recent common ancestor. Furthermore, although these genes are clearly of eubacterial origin, a relationship to the ALS genes of red algae and cyanobacteria (endosymbiotic precursors of plastids) is only weakly indicated. The algal ALS genes are distinguished from their homologs in higher plants by the fact that they are interrupted by numerous spliceosomal introns; plant ALS genes completely lack introns. The restricted phylogenetic distribution of these introns suggests that they were inserted recently, after the divergence of these green algae from plants. Two introns in the Volvox ALS gene, not found in the Chlamydomonas gene, are positioned precisely at sites which resemble "proto-splice" sequences in the Chlamydomonas gene.

 

Kobayashi K, Koyanagi R, Matsumoto M, et al.

Switching from asexual to sexual reproduction in the planarian Dugesia ryukyuensis: Bioassay system and basic description of sexualizing process
ZOOL SCI 16 (2): 291-298 APR 1999

 

Abstract:
An assay system has been established for the sexual induction in the OH strain, an exclusively fissiparous (asexual) strain, of Dugesia ryukyuensis by feeding them with sexually matured worms of Bdellocephala brunnea, an exclusively oviparous (sexual) species. In this assay system, asexual worms gradually differentiated sexual organs, namely the ovary, testis, genital pore and yolk gland in this order, and eventually mated and laid cocoons filled with fertilized eggs. Although the OH strain worms were believed not to have any sexual organs, a pair of undeveloped ovaries with a few oogonia were detected by an intensive histological search. Along with the progression of sexualization, five distinct stages were histologically recognized: In the first stage, the ovaries became larger enough to be externally apparent; oocytes appeared first at stage 2; the primordial testes emerged at stage 3; a genital pore opened, yolk gland primordia developed and spermatocytes appeared at stage 4; and finally at stage 5 matured spermatozoa and yolk glands were formed. Worms in stages 1 and 2 but not in later stages returned asexual if feeding on B, brunnea was interrupted. Furthermore, when the worms at stage 3 onwards were cut posterior to the ovaries, all the tail regenerants developed eventually into fully sexualized worms. Taking these results in account, we have concluded that the process of sexualization has a point-of-no-return between stages 2 and 3. It is likely also that the testes, even the primordia, play an important role in the maintenance and development of sexuality.

 

Hoops HJ, Brighton MC, Stickles SM, et al.

A test of two possible mechanisms for phototactic steering in Volvox carteri (Chlorophyceae)
J PHYCOL 35 (3): 539-547 JUN 1999

Abstract:
We tested two competing models that could explain how differential flagellar activity leads to phototactic turning in spheroids of Volvox carteri f, weismannia (Powers) Iyengar. In one model, turning results from the flagella of anterior cells in the lighted and shadowed hemispheres beating at different frequencies. In a competing model, turning results from a change in beat direction in these flagella. Both models successfully explain phototactic steering under constant illumination, but they make different predictions when colonies are exposed to abrupt changes in light intensity. If turning is due to control of flagellar beat frequency, both progression and rotation rates will change in the same direction and with similar magnitudes. If spheroid turning is due to a change in flagellar beat direction, a decreased rate of progression will accompany an increased rate of rotation and vice versa. We used video-microscopy to observe the behavior of positively phototactic V. carteri spheroids exposed to 10x step-up and step-down stimuli. After a step-up stimulus, spheroids slow their progression and rotation by equal amounts, No significant changes are reported in these parameters after the reciprocal step-down response. These observations are consistent with the variable flagellar frequency model and inconsistent with the variable flagellar direction model for phototactic turning. Switching the direction of Light stimulus by 180 degrees results in reorientation of positively phototactic spheroids, The kinetics of this reorientation did not precisely match the predictions of either model.

 

Nishii I, Ogihara S

Actomyosin contraction of the posterior hemisphere is required for inversion of the Volvox embryo
DEVELOPMENT 126 (10): 2117-2127 MAY 1999

Abstract:
During inversion of a Volvox embryo, a series of cell shape changes causes the multicellular sheet to bend outward, and propagation of the bend from the anterior to the posterior pole eventually results in an inside-out spherical sheet of cells. We use fluorescent and electron microscopy to study the behavior of the cytoskeleton in cells undergoing shape changes. Microtubules are aligned parallel to the cell's long axis and become elongated in the bend. Myosin and actin filaments are arrayed perinuclearly before inversion. In inversion, actin and myosin are located in a subnuclear position throughout the uninverted region but this localization is gradually lost towards the bend. Actomyosin inhibitors cause enlargement of the embryo. The bend propagation is inhibited halfway and, as a consequence, the posterior hemisphere remains uninverted. The arrested posterior hemisphere will resume and complete inversion even in the presence of an actomyosin inhibitor if the anterior hemisphere is removed microsurgically. We conclude that the principal role of actomyosin in inversion is to cause a compaction of the posterior hemisphere; unless the equatorial diameter of the embryo is reduced in this manner, it is too large to pass through the opening defined by the already-inverted anterior hemisphere.

 

Huey RB, Berrigan D, Gilchrist GW, et al.

Testing the adaptive significance of acclimation: A strong inference approach
AM ZOOL 39 (2): 323-336 APR 1999

 

Abstract:
Acclimation is a common phenotypic response to environmental change. Acclimation is often thought to enhance performance and thus to be adaptive. This view has recently been formalized as the "Beneficial Acclimation Hypothesis" and predicts that individuals acclimated to one environment perform better in that environment than do individuals acclimated to a different environment. Although Beneficial Acclimation is appealing and widely supported, recent studies with E. coli and Drosophila have challenged its general validity. Although these challenges could be dismissed as mere exceptions, they encourage a re-evaluation of the adaptive significance of acclimation. Our philosophical approach differs from that of most previous studies of acclimation, in which the prediction derived from a Beneficial Acclimation perspective (e.g., heat tolerance is positively correlated with acclimation temperature) is tested against the null hypothesis ("single hypothesis approach"). Instead, we follow Huey and Berrigan (1996) in advocating a strong inference approach (sensu Platt, 1964), which recognizes that Beneficial Acclimation is actually one of a set of competing hypotheses that make different predictions as to how developmental temperature influences the thermal sensitivity of adults ("developmental acclimation"). Using this perspective, Huey and Berrigan proposed a factorial experimental design (3 developmental by 3 adult temperatures) designed to discriminate among all competing hypotheses. We now derive a formal statistical model (ANOVA with orthogonal polynomial contrasts) for this experimental design and use it to evaluate simultaneously the relative impact of each competing hypothesis. We then apply this model to several case studies (Drosophila, Volvox, Trichogramma), and we review also a recent study with E. coli. The influence of Beneficial Acclimation is supported (albeit often weakly) in most cases. Nevertheless, other hypotheses (especially the Optimal Developmental Temperature Hypothesis) often have a greater impact. Even so, however, Beneficial Acclimation usually predicts relative performance at extreme test temperatures. We conclude that, although rumors of its death are premature, Beneficial Acclimation cannot be viewed as the dominant expectation, at least with regard to developmental temperature acclimation. Moreover, our findings reinforce the view that a strong inference approach provides a more comprehensive portrait of complex biological responses than do single-hypothesis approaches.

 

Hallmann A

Enzymes in the extracellular matrix of Volvox: an inducible, calcium-dependent phosphatase with a modular composition
J BIOL CHEM 274 (3): 1691-1697 JAN 15 1999

Abstract:
The volvocine algae provide the unique opportunity for exploring development of an extracellular matrix. Volvox is the most advanced member of this family and represents the simplest multicellular organism, with differentiated cells, a complete division of labor, and a complex extracellular matrix, which serves structural and enzymatic functions. In Volvox carteri a glycosylated extracellular phosphatase was identified, which is partially released from the extracellular matrix into the growth medium. The phosphatase is synthesized in response to inorganic phosphate starvation and is strictly to inorganic phosphate starvation and is strictly calcium-dependent. The metalloenzyme has been purified to homogeneity and characterized. Its gene and cDNA have been cloned. Comparisons of genomic and cDNA sequences revealed an extremely intron-rich gene (32 introns), With an apparent molecular mass of 160 kDa the Volvox extracellular phosphatase is the largest phosphatase cloned, with no sequence similarity to any other phosphatase. This enzyme exhibits a modular composition. There are two large domains and a small one. The large domains are highly homologous to each other and therefore most likely originated from gene duplication and fusion. At least one EF-hand motif for calcium binding was identified in this extracellular protein Volvox extracellular phosphatase is the first calcium-dependent extracellular phosphatase to be cloned.

 

 

 

 

Rodriguez H, Haring MA, Beck CF

Molecular characterization of two light-induced, gamete-specific genes from Chlamydomonas reinhardtii that encode hydroxyproline-rich proteins
MOL GEN GENET 261 (2): 267-274 MAR 1999

Abstract:
Gametic differentiation in Chlamydomonas reinhardtii is a two-step process, which is controlled by the sequential action of the two extrinsic signals, nitrogen starvation and blue light. The gamete-specific genes GAS28 and GAS29 are expressed in the late phase of gametogenesis. Their light-induced expression is restricted to cells that have completed the first, nitrogen starvation-activated, phase of differentiation. A comparison of the two genes revealed striking similarities as well as differences. Their most prominent shared feature is an extended sequence homology of over 90% in their 5'-untranslated regions, suggesting a role in translational regulation. GAS28 and GAS29 both encode hydroxyproline-rich proteins (HRGPs) of very similar sizes that exhibit typical features of volvocalean cell wall constituents. GAS28 shows a high degree of homology with the Volvox pherophorin gene family, suggesting a relationship between these genes.

 

Bischoff F, Molendijk A, Rajendrakumar CSV, et al.

GTP binding proteins in plants
CELL MOL LIFE SCI 55 (2): 233-256 FEB 1999

Abstract:
GTP-binding proteins are found in all organisms. They are important switches that cycle between an active and an inactive state, ensuring vectorial flow of information on the expense of guanosine triphosphate (GTP). In this review, we discuss current progress in the molecular characterization and functional analysis of plant genes encoding heterotrimeric and small GTPases. An up-to-date list in eluding all cloned plant GTPase genes is given and a systematic classification is proposed.

 

Kirk MM, Stark K, Miller SM, et al.

regA, a Volvox gene that plays a central role in germ-soma differentiation, encodes a novel regulatory protein
DEVELOPMENT 126 (4): 639-647 Feb 1999

 

Abstract:
Volvox has two cell types: mortal somatic cells and immortal germ cells. Here we describe the transposon-tagging, cloning and characterization of regA, which plays a central role as a master regulatory gene in Volvox germsoma differentiation by suppressing reproductive activities in somatic cells. The 12.5 kb regA transcription unit generates a 6,725 nucleotide mRNA that appears at the beginning of somatic cell differentiation, and that encodes a 111 kDa RegA protein that localizes to the nucleus, and has an unusual abundance of alanine, glutamine and proline, This is a compositional feature shared by functional domains of many 'active' repressors, These findings are consistent with the hypothesis that RegA acts in somatic cells to repress transcription of genes required for growth and reproduction, including 13 genes whose products are required for chloroplast biogenesis.

 

Miller SM, Kirk DL

glsA, a Volvox gene required for asymmetric division and germ cell specification, encodes a chaperone-like protein
DEVELOPMENT 126 (4): 649-658 FEB 1999

Abstract:
The gls genes of Volvox are required for the asymmetric divisions that set apart cells of the germ and somatic lineages during embryogenesis. Here we used transposon tagging to clone glsA, and then showed that it is expressed maximally in asymmetrically dividing embryos, and that it encodes a 748-amino acid protein with two potential protein-binding domains. Site-directed mutagenesis of one of these, the J domain (by which Hsp40-class chaperones bind to and activate specific Hsp70 partners) abolishes the capacity of glsA to rescue mutants. Based on this and other considerations, including the fact that the GlsA protein is associated with the mitotic spindle, we discuss how it might function, in conjunction with an Hsp70-type partner, to shift the division plane in asymmetrically dividing cells.

 

Nozaki H, Ohta N, Takano H, et al.

Reexamination of phylogenetic relationships within the colonial Volvocales (Chlorophyta): An analysis of atpB and rbcL gene sequences
J PHYCOL 35 (1): 104-112 FEB 1999

 

Abstract:
The chloroplast-encoded atpB gene was sequenced from 33 strains representing 28 species of the colonial Volvocales (the Volvocaceae and its relatives) to reexamine phylogenetic relationships as previously deduced by morphological data and rbcL gene sequence data.1128 base pairs in the coding regions of the atpB gene were analyzed by MP, NJ, and ML analyses, Although supported with relatively low bootstrap values (75% and 65% in the NJ and ML analyses, respectively), three anisogamous/oogamous volvocacean genera-Eudorina, Pleodorina, and Volvox, excluding the section Volvox(= Euvolvox, illegitimate name), constituted a large monophyletic group (Eudorina group), Outside the Eudorina group, a robust Lineage composed of three species of Volvox sect, Volvox was resolved as in the rbcL gene trees, rejecting the hypothesis of the previous cladistic analysis based on morphological data that the genus Volvox is monophyletic, In addition, the NJ and ML trees suggested that Eudorina is a non-monophyletic genus as inferred from the morphological data and rbcL gene sequences. Although phylogenetic status of the genus Gonium is ambiguous in the rbcL gene trees and the paraphyly of this genus is resolved in the cladistic analysis based on morphological data, the atpB gene sequence data suggest monophyly of Gonium with relatively low bootstrap values (56-61%) in the NJ and ML trees. On the basis of the combined sequence data (2256 base pairs) from atpB and rbcL genes, Gonium was resolved as a robust monophyletic genus in the NJ and ML trees (with 68-86% bootstrap values), and Eudorina elegans Ehrenberg represented a paraphyletic species positioned most basally within the Eudorina group, However, phylogenetic status and relationships of the families of the colonial Volvocales were still almost ambiguous even in the combined analysis.

 

Braun FJ, Hegemann P

Two light-activated conductances in the eye of the green alga Volvox carteri
BIOPHYS J 76 (3): 1668-1678 MAR 1999

Abstract:
Photoreceptor currents of the multicellular green alga Volvox carteri were analyzed using a dissolver mutant. The photocurrents are restricted to the eyespot region of somatic cells. Photocurrents are detectable from intact cells and excised eyes. The rhodopsin action spectrum suggests that the currents are induced by Volvox rhodopsin. Flash-induced photocurrents are a composition of a fast Ca2+-carried current (P-F) and a slower current (P-S), which is carried by H+. P-F is a high-intensity response that appears with a delay of less than 50 mu s after flash. The stimulus-response curve of its initial rise is fit by a single exponential and parallels the rhodopsin bleaching. These two observations suggest that the responsible channel is closely connected to the rhodopsin, both forming a tight complex. At low flash energies P-S is dominating. The current delay increases up to 10 ms, and the P-S amplitude saturates when only a few percent of the rhodopsin is bleached. The data are in favor of a second signaling system, which includes a signal transducer mediating between rhodopsin and the channel. We present a model of how different modes of signal transduction are accomplished in this alga under different light conditions.

 

Hallmann A, Rappel A

Genetic engineering of the multicellular green alga Volvox: a modified and multiplied bacterial antibiotic resistance gene as a dominant selectable marker
PLANT J 17 (1): 99-109 JAN 1999

 

Abstract:
The green alga Volvox represents the simplest multicellular organism: Volvox is composed of only two cell types, somatic and reproductive. Volvox, therefore, is an attractive model system for studying various aspects of multicellularity. With the biolistic nuclear transformation of Volvox carteri, the powerful molecular genetic manipulation of this organism has been established, but applications have been restricted to an auxotrophic mutant serving as the DNA recipient. Therefore, a dominant selectable marker working in all strains and mutants of this organism is required. Among several gene constructs tested, the most advantageous results were obtained with a chimeric gene composed of the coding sequence of the bacterial ble gene, conferring resistance to the antibiotic zeocin, modified with insertions of two endogenous introns from the Volvox arylsulfatase gene and fused to 5' and 3' untranslated regions from the Volvox beta 2-tubulin gene. In the most suitable plasmid used, the gene dosage was increased 16-fold by a technique that allows exponential multiplication of a DNA fragment. Go-transformation of this plasmid and a non-selectable plasmid allowed the identification of zeocin resistant transformants with nuclear integration of both selectable and non-selectable plasmids. Stable expression of the ble gene and of genes from several non-selectable plasmids is demonstrated. The modified ble gene provides the first dominant marker for transformation of both wild-type and mutant strains of Volvox.

 

Fabry S, Kohler A, Coleman AW

Intraspecies analysis: Comparison of ITS sequence data and gene intron sequence data with breeding data for a worldwide collection of Gonium pectorale
J MOL EVOL 48 (1): 94-101 JAN 1999

Abstract:
The morphologically uniform species Gonium pectorale is a colonial green flagellate of worldwide distribution. The affinities of 25 isolates from 18 sites on five continents were assessed by both DNA sequence comparisons and sexual compatibility. Complete sequences were obtained (i) for the internal transcribed spacer ITS-1 and ITS-2 regions of ribosomal DNA and (ii) for each of three single-copy spliceosomal introns, two in a small G protein and one in the actin gene. ITS sequences appeared to homogenize sufficiently rapidly to behave as a single copy gene. Intron sequence differences between isolates in this species reached nucleotide substitution saturation, while ITS sequences did not. Parsimony and evolutionary distance analysis of the two types of DNA data gave essentially the same tree conformation. By all these criteria, the group of G. pectorale isolates fell into two main clades, A and B. Clade A, with isolates from four continents, was comprised of four subclades of quite closely related isolates, plus one strain of ambiguous affinity. Clade B was comprised of two subclades represented by South African and South American isolates, respectively; thus, only subclades of clade B showed geographical localization. With respect to mating, all isolates except one homothallic strain and one apparently sterile strain fell into either one or the other of two mating types. Pairings in all possible combinations revealed that isolates from the same site formed abundant zygotes, which germinated to produce new, sexually active organisms. Zygotes were also formed in many pairings of other combinations, including crosses of clade A with clade B organisms, but none of the latter produced viable germlings. The ability to mate and produce viable progeny that were themselves capable of sexual reproduction was restricted to members of subclades established on the basis of DNA sequence similarities. Thus, the grades of difference in both nuclear intron sequences and rDNA ITS sequences paralleled those observed in the sexual analysis.

 

Miller SM, Kirk DL

glsA, a Volvox gene required for asymmetric division and germ cell specification, encode a chaperone-like protein that co-localizes with the mitotic spindle
MOL BIOL CELL 10: 2265 Suppl. S NOV 1999