Boule and the Evolutionary Origin of Metazoan Gametogenesis: A Grandpa's Tale

The evolution of sex remains a hotly debated topic in evolutionary biology. In particular, studying the origins of the molecular mechanisms underlying sexual reproduction and gametogenesis (its fundamental component) in multicellular eukaryotes has been difficult due to the rapid divergence of many reproductive proteins, pleiotropy, and by the fact that only a very small number of reproductive proteins specifically involved in reproduction are conserved across lineages. Consequently, during the last decade, many efforts have been put into answering the following question: did gametogenesis evolve independently in different animal lineages or does it share a common evolutionary origin in a single ancestral prototype? Among the various approaches carried out in order to solve this question, the characterization of the evolution of the DAZ gene family holds much promise because these genes encode reproductive proteins that are conserved across a wide range of animal phyla. Within this family, BOULE is of special interest because it represents the most ancestral member of this gene family (the “grandfather” of DAZ). Furthermore, BOULE has attracted most of the attention since it represents an ancient male gametogenic factor with an essential reproductive-exclusive requirement in urbilaterians, constituting a core component of the reproductive prototype. Within this context, the aim of the present work is to provide an up-to-date insight into the studies that lead to the characterization of the DAZ family members and the implications in helping decipher the evolutionary origin of gametogenesis in metazoan animals

The evolutionary differentiation of two histone H2A.Z variants in chordates (H2A.Z-1 and H2A.Z-2) is mediated by a stepwise mutation process that affects three amino acid residues

<p>Abstract</p> <p>Background</p> <p>The histone H2A family encompasses the greatest number of core histone variants of which the replacement variant H2A.Z is currently one of the most heavily studied. No clear mechanism for the functional variability that H2A.Z imparts to chromatin has yet been proposed. While most of the past studies have referred to H2A.Z generically as a single protein, in vertebrates it is a mixture of two protein forms H2A.Z-1 (previously H2A.Z) and H2A.Z-2 (previously H2A.F/Z or H2A.V) that differ by three amino acids.</p> <p>Results</p> <p>We have performed an extensive study on the long-term evolution of H2A.Z across metazoans with special emphasis on the possible selective mechanisms responsible for the differentiation between H2A.Z-1 and H2A.Z-2. Our results reveal a common origin of both forms early in chordate evolution. The evolutionary process responsible for the differentiation involves refined stepwise mutation change within the codons of the three differential residues. This eventually led to differences in the intensity of the selective constraints acting upon the different H2A.Z forms in vertebrates.</p> <p>Conclusion</p> <p>The results presented in this work definitively reveal that the existence of H2A.Z-1 and H2A.Z-2 is not a whim of random genetic drift. Our analyses demonstrate that H2A.Z-2 is not only subject to a strong purifying selection but it is significantly more evolutionarily constrained than H2A.Z-1. Whether or not the evolutionary drift between H2A.Z-1 and H2A.Z-2 has resulted in a functional diversification of these proteins awaits further research. Nevertheless, the present work suggests that in the process of their differently constrained evolutionary pathways, these two forms may have acquired new or complementary functions.</p

Caracterización de los genes de histonas en el género Mytilus y evolución molecular de la familia multigénica H1

[Resumen] INTRODUCCIÓN Las histonas son un grupo de proteínas básicas de pequeño tamaño presentes en las células eucariotas, involucradas en el empaquetamiento del ADN formando la fibra de cromatina. Existen cinco familias que se clasifican en histonas del core (H2A, H2B, H3, H4) e histonas linker (H1). Su origen evolutivo parece remontarse a organismos procariotas, y aunque es obvio que la heterogeneidad funcional en eucariotas debe venir dictada por mecanismos complejos, hasta la fecha su evolución ha sido explicada como un proceso concertado. El conocimiento actual es limitado en cuanto a patrones de organización, expresión y evolución de estos genes en invertebrados, especialmente en el caso de genes H1. Para mejorar el conocimiento acerca de las histonas se han propuesto tres objetivos principales: Caracterizar la unidad repetitiva de histonas en el molusco bivalvo Mytilus galloprovincialis, estudiar las secuencias de ADN de los cinco genes de histonas en cuatro especies adicionales (M. californianus, M. chilensis, M. edulis y M. trossulus), y analizar los mecanismos involucrados en la evolución de los genes H1. RESULTADOS Se diseñaron primers para amplificar mediante PCR la refión codificante de cada una de las cinco histonas en especies del género Mytilus, los cuáles permitieron obtener sondas para buscar genes de histonas en una genoteca de M. galloprovincialis, caracterizándose la unidad repetitiva como H4 mayor que , menor que H2B, H2A mayor que , H3 mayor que , H1 mayor que , 5S mayor que , 5S mayor que . Las regiones promotoras mostraron la presencia de elementos reguladores típicos (cajas TATA, posiciones CAP y cajas CAAT). En el caso de H1 también se identificaron elementos H1 box-like y elementos H4 box, mientras que las regiones 3' terminales UTR mostraron secuencias tallo-bucle, seguidas por un elemento rico en purinas y al menos una señal de poli(A). La caracterización de la unidad en M. galloprovincial

Molecular and evolutionary analysis of mussel histone genes ("Mytilus" spp.): possible evidence of an "orphon origin" for H1 histone genes

[Abstract:] Linker histones are a divergent group of histone proteins with an independent evolutionary history in which, besides somatic subtypes, tissue- and differentiation-specific subtypes are included. In the present work H1 histone coding and noncoding segments from five Mytilus mussel species (Mollusca: Bivalvia) widely distributed throughout the world have been determined and characterized. Analysis of promoter regions shows clear homologies among Mytilus H1 genes, sea urchin H1 genes, and vertebrate differentiation-specific H1 subtypes (H5 and H10), all having an H4 box motif in common. The amino acid sequence of the H1 protein central conserved domain is also closely related to that previously defined for the vertebrate divergent subtypes. A phylogenetic tree reconstructed from different H1 genes from several species strengthens the hypothesis of an “orphon” origin for the Mytilus H1 genes, as well as for the H10/H5 genes from vertebrates and the H1D gene from the sea urchin Strongylocentrotus purpuratus, is suggested. As additional data, the average copy number of the H1 genes in the species analyzed was estimated as being 100 to 110 copies per haploid genome, where FISH revealed telomeric chromosomal location for several H1 copies in M. galloprovincialis. The contribution of such proximity to heterochromatic regions over the amount of codon bias detected for H1 genes is discussed.Ministerio de Ciencia e Innovación; IFD97-129

Birth-and-death evolution with strong purifying selection in the histone H1 multigene family and the origin of "orphon" H1 genes

[Abstract:] Histones are small basic nuclear proteins with critical structural and functional roles in eukaryotic genomes. The H1 multigene family constitutes a very interesting histone class gathering the greatest number of isoforms, with many different arrangements in the genome, including clustered and solitary genes, and showing replication-dependent (RD) or replication-independent (RI) expression patterns. The evolution of H1 histones has been classically explained by concerted evolution through a rapid process of interlocus recombination or gene conversion. Given such intriguing features, we have analyzed the long-term evolutionary pattern of the H1 multigene family through the evaluation of the relative importance of gene conversion, point mutation, and selection in generating and maintaining the different H1 subtypes. We have found the presence of an extensive silent nucleotide divergence, both within and between species, which is always significantly greater than the nonsilent variation, indicating that purifying selection is the major factor maintaining H1 protein homogeneity. The results obtained from phylogenetic analysis reveal that different H1 subtypes are no more closely related within than between species, as they cluster by type in the topologies, and that both RD and RI H1 variants follow the same evolutionary pattern. These findings suggest that H1 histones have not been subject to any significant effect of interlocus recombination or concerted evolution. However, the diversification of the H1 isoforms seems to be enhanced primarily by mutation and selection, where genes are subject to birth-and-death evolution with strong purifying selection at the protein level. This model is able to explain not only the generation and diversification of RD H1 isoforms but also the origin and long-term persistence of orphon RI H1 subtypes in the genome, something that is still unclear, assuming concerted evolution.Xunta de Galicia; PGIDT (10PX110304

The characterization of amphibian nucleoplasmins yields new insight into their role in sperm chromatin remodeling

BACKGROUND: Nucleoplasmin is a nuclear chaperone protein that has been shown to participate in the remodeling of sperm chromatin immediately after fertilization by displacing highly specialized sperm nuclear basic proteins (SNBPs), such as protamine (P type) and protamine-like (PL type) proteins, from the sperm chromatin and by the transfer of histone H2A-H2B. The presence of SNBPs of the histone type (H type) in some organisms (very similar to the histones found in somatic tissues) raises uncertainty about the need for a nucleoplasmin-mediated removal process in such cases and poses a very interesting question regarding the appearance and further differentiation of the sperm chromatin remodeling function of nucleoplasmin and the implicit relationship with SNBP diversity The amphibians represent an unique opportunity to address this issue as they contain genera with SNBPs representative of each of the three main types: Rana (H type); Xenopus (PL type) and Bufo (P type). RESULTS: In this work, the presence of nucleoplasmin in oocyte extracts from these three organisms has been assessed using Western Blotting. We have used mass spectrometry and cloning techniques to characterize the full-length cDNA sequences of Rana catesbeiana and Bufo marinus nucleoplasmin. Northern dot blot analysis shows that nucleoplasmin is mainly transcribed in the egg of the former species. Phylogenetic analysis of nucleoplasmin family members from various metazoans suggests that amphibian nucleoplasmins group closely with mammalian NPM2 proteins. CONCLUSION: We have shown that these organisms, in striking contrast to their SNBPs, all contain nucleoplasmins with very similar primary structures. This result has important implications as it suggests that nucleoplasmin's role in chromatin assembly during early zygote development could have been complemented by the acquisition of a new function of non-specifically removing SNBPs in sperm chromatin remodeling. This acquired function would have been strongly determined by the constraints imposed by the appearance and differentiation of SNBPs in the sperm

Histone H2A (H2A.X and H2A.Z) Variants in Molluscs: Molecular Characterization and Potential Implications For Chromatin Dynamics

Histone variants are used by the cell to build specialized nucleosomes, replacing canonical histones and generating functionally specialized chromatin domains. Among many other processes, the specialization imparted by histone H2A (H2A.X and H2A.Z) variants to the nucleosome core particle constitutes the earliest response to DNA damage in the cell. Consequently, chromatin-based genotoxicity tests have been developed in those cases where enough information pertaining chromatin structure and dynamics is available (i.e., human and mouse). However, detailed chromatin knowledge is almost absent in most organisms, specially protostome animals. Molluscs (which represent sentinel organisms for the study of pollution) are not an exception to this lack of knowledge. In the present work we first identified the existence of functionally differentiated histone H2A.X and H2A.Z variants in the mussel Mytilus galloprovincialis (MgH2A.X and MgH2A.Z), a marine organism widely used in biomonitoring programs. Our results support the functional specialization of these variants based on: a) their active expression in different tissues, as revealed by the isolation of native MgH2A.X and MgH2A.Z proteins in gonad and hepatopancreas; b) the evolutionary conservation of different residues encompassing functional relevance; and c) their ability to confer specialization to nucleosomes, as revealed by nucleosome reconstitution experiments using recombinant MgH2A.X and MgH2A.Z histones. Given the seminal role of these variants in maintaining genomic integrity and regulating gene expression, their preliminary characterization opens up new potential applications for the future development of chromatin-based genotoxicity tests in pollution biomonitoring programs

The gene transformer-2 of Anastrepha fruit flies (Diptera, Tephritidae) and its evolution in insects

<p>Abstract</p> <p>Background</p> <p>In the tephritids <it>Ceratitis</it>, <it>Bactrocera </it>and <it>Anastrepha</it>, the gene <it>transformer </it>provides the memory device for sex determination via its auto-regulation; only in females is functional Tra protein produced. To date, the isolation and characterisation of the gene <it>transformer-2 </it>in the tephritids has only been undertaken in <it>Ceratitis</it>, and it has been shown that its function is required for the female-specific splicing of <it>doublesex </it>and <it>transformer </it>pre-mRNA. It therefore participates in <it>transformer </it>auto-regulatory function. In this work, the characterisation of this gene in eleven tephritid species belonging to the less extensively analysed genus <it>Anastrepha </it>was undertaken in order to throw light on the evolution of <it>transformer-2</it>.</p> <p>Results</p> <p>The gene <it>transformer-2 </it>produces a protein of 249 amino acids in both sexes, which shows the features of the SR protein family. No significant partially spliced mRNA isoform specific to the male germ line was detected, unlike in <it>Drosophila</it>. It is transcribed in both sexes during development and in adult life, in both the soma and germ line. The injection of <it>Anastrepha transformer-2 </it>dsRNA into <it>Anastrepha </it>embryos caused a change in the splicing pattern of the endogenous <it>transformer </it>and <it>doublesex </it>pre-mRNA of XX females from the female to the male mode. Consequently, these XX females were transformed into pseudomales. The comparison of the eleven <it>Anastrepha </it>Transformer-2 proteins among themselves, and with the Transformer-2 proteins of other insects, suggests the existence of negative selection acting at the protein level to maintain Transformer-2 structural features.</p> <p>Conclusions</p> <p>These results indicate that <it>transformer-2 </it>is required for sex determination in <it>Anastrepha </it>through its participation in the female-specific splicing of <it>transformer </it>and <it>doublesex </it>pre-mRNAs. It is therefore needed for the auto-regulation of the gene <it>transformer</it>. Thus, the <it>transformer/transfomer-2 > doublesex </it>elements at the bottom of the cascade, and their relationships, probably represent the ancestral state (which still exists in the Tephritidae, Calliphoridae and Muscidae lineages) of the extant cascade found in the Drosophilidae lineage (in which <it>tra </it>is just another component of the sex determination gene cascade regulated by <it>Sex-lethal</it>). In the phylogenetic lineage that gave rise to the drosophilids, evolution co-opted for <it>Sex-lethal</it>, modified it, and converted it into the key gene controlling sex determination.</p

Common evolutionary origin and birth-and-death process in the replication-independent histone H1 isoforms from vertebrate and invertebrate genomes

[Abstract]The H1 histone multigene family shows the greatest diversity of isoforms among the five histone gene families, including replication-dependent (RD) and replication-independent (RI) genes, according to their expression patterns along the cell cycle and their genomic organization. Although the molecular characterization of the RI isoforms has been well documented in vertebrates, similar information is lacking in invertebrates. In this work we provide evidence for a polyadenylation signature in the Mytilus “orphon” H1 genes similar to the polyadenylation characteristic of RI H1 genes. These mussel genes, together with the sea urchin H1δ genes, are part of a lineage of invertebrate “orphon” H1 genes that share several control elements with vertebrate RI H1 genes. These control elements include the UCE element, H1-box and H4-box. We provide evidence for a functional evolution of vertebrate and invertebrate RI H1 genes, which exhibit a clustering pattern by type instead of by species, with a marked difference from the somatic variants. In addition, these genes display an extensive silent divergence at the nucleotide level which is always significantly larger than the nonsilent. It thus appears that RI and RD H1 isoforms display similar long-term evolutionary patterns, best described by the birth-and-death model of evolution. Notably, this observation is in contrast with the theoretical belief that clustered RD H1 genes evolve in a concerted manner. The split of the RI group from the main RD group must therefore have occurred before the divergence between vertebrates and invertebrates about 815 million years ago. This was the result of the transposition of H1 genes to solitary locations in the genome.Xunta de Galicia; 10PX110304Canadá. Canadian Institutes of Health Research; MOP-5771