A new deletion of the mouse Y chromosome long arm associated with the loss of Ssty expression, abnormal sperm development and sterility

The mouse Y chromosome carries 10 distinct genes or gene families that have open reading frames suggestive of retained functionality; it has been assumed that many of these function in spermatogenesis. However, we have recently shown that only two Y genes, the testis determinant Sry and the translation initiation factor Eif2s3y, are essential for spermatogenesis to proceed to the round spermatid stage. Thus, any further substantive mouse Y-gene functions in spermatogenesis are likely to be during sperm differentiation. The complex Ssty gene family present on the mouse Y long arm (Yq) has been implicated in sperm development, with partial Yq deletions that reduce Ssty expression resulting in impaired fertilization efficiency. Here we report the identification of a more extensive Yq deletion that abolishes Ssty expression and results in severe sperm defects and sterility. This result establishes that genetic information (Ssty?) essential for normal sperm differentiation and function is present on mouse Yq.     

Yeast Srp1, a nuclear protein related toDrosophila and mouse pendulin, is required for normal migration, division, and integrity of nuclei during mitosis

This paper describes genes from yeast and mouse with significant sequence similarities to aDrosophila gene that encodes the blood cell tumor suppressor pendulin. The protein encoded by the yeast gene, Srp1p, and mouse pendulin share 42% and 51% amino acid identity withDrosophila pendulin, respectively. All three proteins consist of 10.5 degenerate tandem repeats of 42 amino acids each. Similar repeats occur in a superfamily of proteins that includes theDrosophila Armadillo protein. All three proteins contain a consensus sequence for a bipartite nuclear localization signal (NLS) in the N-terminal domain, which is not part of the repeat structure. Confocal microscopic analysis of yeast cells stained with antibodies against Srp1p reveals that this protein is intranuclear throughout the cell cycle. Targeted gene disruption shows thatSRP1 is an essential gene. Despite their sequence similarities,Drosophila and mouse pendulin are unable to rescue the lethality of anSRP1 disruption. We demonstrate that yeast cells depleted of Srp1p arrest in mitosis with a G2 content of DNA. Arrested cells display abnormal structures and orientations of the mitotic spindles, aberrant segregation of the chromatin and the nuclei, and threads of chromatin emanating from the bulk of nuclear DNA. This phenotype suggests that Srplp is required for the normal function of microtubules and the spindle pole bodies, as well as for nuclear integrity. We suggest that Srp1p interacts with multiple components of the cell nucleus that are required for mitosis and discuss its functional similarities to, and differences fromDrosophila pendulin.     

Comparative chloroplast genomes of Paris Sect. Marmorata: insights into repeat regions and evolutionary implications

Background

Species of Paris Sect. Marmorata are valuable medicinal plants to synthesize steroidal saponins with effective pharmacological therapy. However, the wild resources of the species are threatened by plundering exploitation before the molecular genetics studies uncover the genomes and evolutionary significance. Thus, the availability of complete chloroplast genome sequences of Sect. Marmorata is necessary and crucial to the understanding the plastome evolution of this section and facilitating future population genetics studies. Here, we determined chloroplast genomes of Sect. Marmorata, and conducted the whole chloroplast genome comparison.

Results

This study presented detailed sequences and structural variations of chloroplast genomes of Sect. Marmorata. Over 40 large repeats and approximately 130 simple sequence repeats as well as a group of genomic hotspots were detected. Inverted repeat contraction of this section was inferred via comparing the chloroplast genomes with the one of P. verticillata. Additionally, almost all the plastid protein coding genes were found to prefer ending with A/U. Mutation bias and selection pressure predominately shaped the codon bias of most genes. And most of the genes underwent purifying selection, whereas photosynthetic genes experienced a relatively relaxed purifying selection.

Conclusions

Repeat sequences and hotspot regions can be scanned to detect the intraspecific and interspecific variability, and selected to infer the phylogenetic relationships of Sect. Marmorata and other species in subgenus Daiswa. Mutation and natural selection were the main forces to drive the codon bias pattern of most plastid protein coding genes. Therefore, this study enhances the understanding about evolution of Sect. Marmorata from the chloroplast genome, and provide genomic insights into genetic analyses of Sect. Marmorata.

     

Identification of novel Y chromosome encoded transcripts by testis transcriptome analysis of mice with deletions of the Y chromosome long arm

Background  

The male-specific region of the mouse Y chromosome long arm (MSYq) is comprised largely of repeated DNA, including multiple copies of the spermatid-expressed Ssty gene family. Large deletions of MSYq are associated with sperm head defects for which Ssty deficiency has been presumed to be responsible.     

Uno studio citofotometrico delle proteine durante la mitosi nel meristema radicale di Allium cepa L.

Abstract

A cytophotometric study of proteins during mitosis in root tip meristematic cells of «Allium cepa L.». — Spectrophotometric analyses of the amount of Fast-green stainable proteins at different pH values (8,1; 6; 4; 2) have been accomplished in Allium cepa root tip meristematic cells during the four phases of mitosis. The results seem to indicate that: a) the highest absorption is detectable in correspondence of metaphase at each of the four pH values; b) the transition from prophase to metaphase is characterized by an increase of both proteins reacting at pH 8,1 (histones) and between pH 6 and pH 8,1 (neutral proteins); c) the transition from metaphase to anaphase is characterized by a loss of histones and of proteins reacting between pH 4 and pH 6 (acid proteins); d) the transition from anaphase to telophase is characterized by a loss of neutral proteins. The data are discussed in relation to the problem of chromosome coiling.     

Domain structure and conserved epitopes of Sfb protein, the fibronectin-binding adhesin of Streptococcus pyogenes

Streptococcus pyogenes expresses a fibronectin-binding surface protein (Sfb protein) which mediates adherence to human epithelial cells. The nucleotide sequence of the sfb gene was determined and the primary sequence of the Sfb protein was analysed. The protein consists of 638 amino acids and comprises five structurally distinct domains. The protein starts with an N-terminal signal peptide followed by an aromatic domain. The central part of the protein is formed by four proline-rich repeats which are flanked by non-repetitive spacer sequences. A second repeat region, consisting of four repeats that are distinct from the proline repeats and have been shown to form the fibronectin-binding domain, is located in the Cterminal part of the protein. The protein ends with a typical cell wall and membrane anchor region. Comparative sequence analysis of the N-terminal aromatic domain revealed similarities with carbohydrate-binding sites of other proteins. The proline repeat region of the Sfb protein shares characteristic features with proline-rich repeats of functionally distinct surface proteins from pathogenic Gram-positive cocci. Immunoelectron microscopy revealed an even distribution of the fibronectin-binding domain of Sfb protein on the surface of streptococcal cells. Analyses of 38 sfb genes originating from different S. pyogenes isolates revealed primary sequence variability in regions coding for the N-termini of mature Sfb proteins, whereas sequences coding for the central and C-terminal repeats were highly conserved. The repeat sequences are postulated to act as target sites for intragenic recombination events that result in variable numbers of repeats within the different sfb genes. A model of the Sfb protein is presented.     

Computational Approaches and Tools Used in Identification of Dispersed Repetitive DNA Sequences

It has become clear that dispersed repeat sequences have played multiple roles in eukaryotic genome evolution including increasing genetic diversity through mutation, inducing changes in gene expression, and facilitating generation of novel genes. Growing recognition of the importance of dispersed repeats has fueled development of computational tools designed to expedite discovery and classification of repeats. Here we review major existing repeat exploration tools and discuss the algorithms utilized by these tools. Special attention is devoted to ab initio programs, i.e., those tools that do not rely upon previously identified repeats to find new repeat elements. We conclude by discussing the strengths and weaknesses of current tools and highlighting additional approaches that may advance repeat discovery/characterization.     

Analysis of putative DNA amplification genes in the element AUD1 of Streptomyces lividans 66

The amplifiable AUD1 element of Streptomyces lividans 66 consists of two copies of a 4.7 kb sequence flanked by three copies of a 1 kb sequence. The DNA sequences of the three 1 kb repeats were determined. Two copies (left and middle repeats) were identical: (1009 by in length) and the right repeat was 1012 bp long and differed at 63 positions. The repeats code for open reading frames (ORFs) with typical Streptomyces codon usage, which would encode proteins of about 36 kD molecular weight. The sequences of these ORFs suggest that they specify DNA-binding proteins and potential palindromic binding sites are found adjacent to the genes. The putative amplification protein encoded by the right repeat was expressed in Escherichia coli.     

The four repeat Giardia lamblia telomere forms tetramolecular G-quadruplex with antiparallel topology

Abstract

Guanine rich DNA sequences of regulatory genomic regions form secondary structures known as G-quadruplexes usually stabilized by tetrads of Hoogsteen hydrogen bonded guanines. The in vivo existence of G-quadruplexes ascertains their biological roles. Human telomeric repeats are the most studied G-rich sequences. The four repeat Giardia telomeric sequence (TAGGG)₄ differs from its human counterpart (TTAGGG)_4, by deletion of one T at the G-tract intervening site of each repeat. We show here that whilst the two repeat Giardia telomeric sequence (TAGGG)₂ forms parallel and antiparallel quadruplexes with tetramolecular topology exclusively, the four repeat version (TAGGG)₄ forms a tetramolecular (antiparallel) and unimolecular (parallel) quadruplexes in Na⁺. The tetramolecular (antiparallel) G-quadruplex formed by four repeats of Giardia telomeric sequence is stabilized by the additional Watson-Crick bonding between its intervening TA bases aligned in antiparallel fashion. Four stranded antiparallel quadruplex for four repeats of any telomeric sequence have not been characterized till date. We hypothesize that telomeric association in antiparallel fashion, (via G-overhangs to form tetramolecular quadruplex) could be a biologically relevant molecular event. Further, coexistence of Hoogsteen as well as Watson-Crick base pairing might give insight for recognition of conformationally diverse DNA structures by ligands.

Communicated by Ramaswamy H. Sarma     

Bidirectional transcription of a novel chimeric gene mapping to mouse chromosome Yq

Background  

The male-specific region of the mouse Y chromosome long arm (MSYq) contains three known highly multi-copy X-Y homologous gene families, Ssty1/2, Sly and Asty. Deletions on MSYq lead to teratozoospermia and subfertility or infertility, with a sex ratio skew in the offspring of subfertile MSYqdel males     

A protein encoded by a member of the multicopy Ssty gene family located on the long arm of the mouse Y chromosome is expressed during sperm development

Multicopy Y-chromosomal genes in human and mouse have been postulated to play a role in spermatogenesis. The mouse Y long arm (Yq) carries hundreds of supposedly intronless copies of Ssty, for which no protein has hitherto been identified; mice lacking Yq are sterile with grossly abnormal sperm. We have now identified an Ssty-encoded protein (Ssty1) that is expressed in spermatids. The protein is absent from spermatids of mice that lack Yq, but is not reduced in mice with a two-thirds reduction of Ssty copies, implying that most do not produce this protein. Furthermore, no protein was produced by a strongly transcribed intronless Ssty transgene, raising doubts as to the protein-encoding potential of these intronless genes. We have now identified an intron-containing copy that is also present in multiple copies on Yq. One or more intron-containing copies are retained in the Ssty-deficient mice and may be the source of the Ssty1 protein.     

斑马鱼DrSpin-1基因的克隆和特征分析

Spin蛋白家族是具有Spin/Ssty保守结构域并在配子发生过程中发挥关键作用的一类分子。研究利用简并引物PCR,从斑马鱼成熟卵母细胞SMART cDNA文库中筛选到260 bp的DrSpin-1和DrSpin-2部分序列,经序列同源性比对,斑马鱼DrSpin-1的部分氨基酸序列与银鲫CagSpin一致性高达81%。利用RACEPCR从该cDNA文库中获得斑马鱼DrSpin-1的全长cDNA序列。序列分析表明,DrSpin-1全长cDNA为1082 bp,开放阅读框771 bp,编码257个氨基酸,具有三个Spin/Ssty保守域,8个可能的磷酸化位点,初步确定斑马鱼DrSpin-1是Spin基因家族成员。斑马鱼DrSpin-1蛋白与已报道的鱼类Spin蛋白多重序列比对表明,DrSpin-1蛋白与银鲫CagSpin蛋白同源性最高。可以推测克隆得到的斑马鱼DrSpin-1与已知功能的银鲫CagSpin具有相近的表达谱和生物学功能,可能在配子发生和受精过程中发挥重要作用。     

Comparative Analysis of the Proteins with Tandem Repeats from 8 Microsporidia and Characterization of a Novel Endospore Wall Protein Colocalizing with Polar Tube from Nosema bombycis

As a common feature of eukaryotic proteins, tandem amino acid repeat has been studied extensively in both animal and plant proteins. Here, a comparative analysis focusing on the proteins having tandem repeats was conducted in eight microsporidia, including four mammal‐infecting microsporidia (Encephalitozoon cuniculi, Encephalitozoon intestinalis, Encephalitozoon hellem and Encephalitozoon bieneusi) and four insect‐infecting microsporidia (Nosema apis, Nosema ceranae, Vavraia culicis and Nosema bombycis). We found that the proteins with tandem repeats were abundant in these species. The quantity of these proteins in insect‐infecting microsporidia was larger than that of mammal‐infecting microsporidia. Additionally, the hydrophilic residues were overrepresented in the tandem repeats of these eight microsporidian proteins and the amino acids residues in these tandem repeat sequences tend to be encoded by GC‐rich codons. The tandem repeat position within proteins of insect‐infecting microsporidia was randomly distributed, whereas the tandem repeats within proteins of mammal‐infecting microsporidia rarely tend to be present in the N terminal regions, when compared with those present in the C terminal and middle regions. Finally, a hypothetical protein EOB14572 possessing four tandem repeats was successfully characterized as a novel endospore wall protein, which colocalized with polar tube of N. bombycis. Our study provided useful insight for the study of the proteins with tandem repeats in N. bombycis, but also further enriched the spore wall components of this obligate unicellular eukaryotic parasite.     

The Predicted Presence of Large Helical Structural Variation in Yeast HIS4 Upstream Region is Correlated with General Amino Acid Control on the CYC1 Gene

Abstract

A series of CYC1 constructions in which the upstream promoter portion has been replaced by a variety of HIS4 synthetic fragments has demonstrated that the 5′ TGACTC 3′ repeat is crucial for conferring amino acid general control. Efficient regulation, however, is obtained only with fragments containing both the repeat and flanking sequences. Analysis of the flanks shows the presence of a 16 nucleotide long sequence composed of alternations of two purines and two pyrimidines between the upstream and downstream repeats. Such a sequence has very large twist angle variations. Homologous sequences are observed in HIS1, HIS3, and in TRP5 upstream regions between copies of the repeat. Sequences which confer special structural characteristics may aid in protein recognition of the promoter region.