3640 Unique EST Clusters from the Medaka Testis and Their Potential Use for Identifying Conserved Testicular Gene Expression in Fish and Mammals

Background

The fish medaka is the first vertebrate capable of full spermatogenesis in vitro from self-renewing spermatogonial stem cells to motile test-tube sperm. Precise staging and molecular dissection of this process has been hampered by the lack of suitable molecular markers.

Methodology and Principal Findings

We have generated a normalized medaka testis cDNA library and obtained 7040 high quality sequences representing 3641 unique gene clusters. Among these, 1197 unique clusters are homologous to known genes, and 2444 appear to be novel genes. Ontology analysis shows that the 1197 gene products are implicated in diverse molecular and cellular processes. These genes include markers for all major types of testicular somatic and germ cells. Furthermore, markers were identified for major spermatogenic stages ranging from spermatogonial stem cell self-renewal to meiosis entry, progression and completion. Intriguingly, the medaka testis expresses at least 13 homologs of the 33 mouse X-chromosomal genes that are enriched in the testis. More importantly, we show that key components of several signaling pathways known to be important for testicular function in mammals are well represented in the medaka testicular EST collection.

Conclusions/Significance

Medaka exhibits a considerable similarity in testicular gene expression to mammals. The medaka testicular EST collection we obtained has wide range coverage and will not only consolidate our knowledge on the comparative analysis of known genes'' functions in the testis but also provide a rich resource to dissect molecular events and mechanism of spermatogenesis in vivo and in vitro in medaka as an excellent vertebrate model.     

Genomic analysis of the relationship between gene expression variation and DNA polymorphism in Drosophila simulans

Background

Understanding how DNA sequence polymorphism relates to variation in gene expression is essential to connecting genotypic differences with phenotypic differences among individuals. Addressing this question requires linking population genomic data with gene expression variation.

Results

Using whole genome expression data and recent light shotgun genome sequencing of six Drosophila simulans genotypes, we assessed the relationship between expression variation in males and females and nucleotide polymorphism across thousands of loci. By examining sequence polymorphism in gene features, such as untranslated regions and introns, we find that genes showing greater variation in gene expression between genotypes also have higher levels of sequence polymorphism in many gene features. Accordingly, X-linked genes, which have lower sequence polymorphism levels than autosomal genes, also show less expression variation than autosomal genes. We also find that sex-specifically expressed genes show higher local levels of polymorphism and divergence than both sex-biased and unbiased genes, and that they appear to have simpler regulatory regions.

Conclusion

The gene-feature-based analyses and the X-to-autosome comparisons suggest that sequence polymorphism in cis-acting elements is an important determinant of expression variation. However, this relationship varies among the different categories of sex-biased expression, and trans factors might contribute more to male-specific gene expression than cis effects. Our analysis of sex-specific gene expression also shows that female-specific genes have been overlooked in analyses that only point to male-biased genes as having unusual patterns of evolution and that studies of sexually dimorphic traits need to recognize that the relationship between genetic and expression variation at these traits is different from the genome as a whole.     

Evolutionary rate and gene expression across different brain regions

Background

The evolutionary rate of a protein is a basic measure of evolution at the molecular level. Previous studies have shown that genes expressed in the brain have significantly lower evolutionary rates than those expressed in somatic tissues.

Results

We study the evolutionary rates of genes expressed in 21 different human brain regions. We find that genes highly expressed in the more recent cortical regions of the brain have lower evolutionary rates than genes highly expressed in subcortical regions. This may partially result from the observation that genes that are highly expressed in cortical regions tend to be highly expressed in subcortical regions, and thus their evolution faces a richer set of functional constraints. The frequency of mammal-specific and primate-specific genes is higher in the highly expressed gene sets of subcortical brain regions than in those of cortical brain regions. The basic inverse correlation between evolutionary rate and gene expression is significantly stronger in brain versus nonbrain tissues, and in cortical versus subcortical regions. Extending upon this cortical/subcortical trend, this inverse correlation is generally more marked for tissues that are located higher along the cranial vertical axis during development, giving rise to the possibility that these tissues are also more evolutionarily recent.

Conclusions

We find that cortically expressed genes are more conserved than subcortical ones, and that gene expression levels exert stronger constraints on sequence evolution in cortical versus subcortical regions. Taken together, these findings suggest that cortically expressed genes are under stronger selective pressure than subcortically expressed genes.     

Induction of diploid gynogenesis in an evolutionary model organism, the three-spined stickleback (Gasterosteus aculeatus)

Background

X-linked alpha thalassemia, mental retardation syndrome in humans is a rare recessive disorder caused by mutations in the ATRX gene. The disease is characterised by severe mental retardation, mild alpha-thalassemia, microcephaly, short stature, facial, skeletal, genital and gonadal abnormalities.

Results

We examined the expression of ATRX and ATRY during early development and gonadogenesis in two distantly related mammals: the tammar wallaby (a marsupial) and the mouse (a eutherian). This is the first examination of ATRX and ATRY in the developing mammalian gonad and fetus. ATRX and ATRY were strongly expressed in the developing male and female gonad respectively, of both species. In testes, ATRY expression was detected in the Sertoli cells, germ cells and some interstitial cells. In the developing ovaries, ATRX was initially restricted to the germ cells, but was present in the granulosa cells of mature ovaries from the primary follicle stage onwards and in the corpus luteum. ATRX mRNA expression was also examined outside the gonad in both mouse and tammar wallaby whole embryos. ATRX was detected in the developing limbs, craniofacial elements, neural tissues, tail and phallus. These sites correspond with developmental deficiencies displayed by ATR-X patients.

Conclusions

There is a complex expression pattern throughout development in both mammals, consistent with many of the observed ATR-X syndrome phenotypes in humans. The distribution of ATRX mRNA and protein in the gonads was highly conserved between the tammar and the mouse. The expression profile within the germ cells and somatic cells strikingly overlaps with that of DMRT1, suggesting a possible link between these two genes in gonadal development. Taken together, these data suggest that ATRX has a critical and conserved role in normal development of the testis and ovary in both the somatic and germ cells, and that its broad roles in early mammalian development and gonadal function have remained unchanged for over 148 million years of mammalian evolution.     

Intérêt de l’abaissement du testicule non descendu chez l’adulte. Etude à propos de 259 patients

Introduction

Undescended testis is a frequent congenital disease, more often diagnosed and treated during childhood. However, due to ignorance or negligence, this disease can be seen even after puberty, when it raises a therapeutic problem: is orchidopexy still useful? This study was designed to evaluate the outcome of orchidopexy at adulthood in terms of improvement of fertility and prevention of malignant degeneration.

Material and methods

Retrospective study performed over a 23-year period (1983–2005). We have found 259 patients with undescended testis diagnosed and treated after the age of 18 years.

Results

The mean age of patients was 24 years (range: 18–63). In the majority of cases, undescended testis was diagnosed at a systemic medical examination in 199 patients (77% of cases), and in a context of infertility in 33 patients, testicular malignancy in 8 patients, testicular torsion in 2 patients and, in 17 cases, undescended testis had been known since birth but was neglected by the parents. Undescended testis was unilateral in 209 cases and bilateral in 50 cases. Out of 37 couples, only 4 gave birth to children (10.8% paternity rate). Sperm analysis was abnormal in all infertile patients. All patients were treated by orchidopexy, except for 36 patients in whom orchidectomy was performed due to testicular atrophy (27 cases), malignancy (8 cases) or necrosis (1 case). Biopsy of the intrascrotal testis was performed in 3 patients with unilateral cryptorchidism. Histological examination was normal in two cases and abnormal in one case. The long-term outcome was characterized by:

1. Testicular atrophy in 6 patients (2.7% of cases).
2. Progression to malignancy in 3 patients (1.3% of cases).
3. Improvement of sperm parameters in 16 of 33 infertile patients (48.5%); 4 patients fathered children after treatment.

Conclusion

Orchidopexy at adulthood can lead to improvement of infertility. It can also decrease the incidence of malignancy and facilitate clinical examination looking for possible malignancy. However, the best treatment remains preventive, based on early diagnosis and orchidopexy.     

Genetic correlates of autoreactivity and autoreactive potential in human Ig heavy chains

Background

Immature bone marrow B cells are known to have longer CDR3 than mature peripheral B cells, and this genetic characteristic has been shown to correlate with autoreactivity in these early cells. B-cell Central tolerance eliminates these cells, but it is known that autoreactive B cells nevertheless appear commonly in healthy human blood. We examined over 7,300 Ig genes from Genbank, including those annotated by their discoverers as associated with autoreactivity, to determine the genetic correlates of autoreactivity in mature B cells.

Results

We find differential biases in gene segment usage and higher mutation frequency in autoreactivity-associated Ig genes, but the CDR3 lengths do not differ between autoreactive and non-autoreactive Ig genes. The most striking genetic signature of autoreactivity is an increase in the proportion of N-nucleotides relative to germline-encoded nucleotides in CDR3 from autoreactive genes.

Conclusion

We hypothesize that peripheral autoreactivity results primarily from somatic mutation, and that the genetic correlates of autoreactivity in mature B-cells are not the same as those for autoreactivity in immature B cells. What is seen in mature autoreactive B cells are the correlates of autoreactive potential, not of autoreactivity per se. The autoreactive potential is higher for V(D)J rearrangements encoded to a large extent by N-nucleotides rather than by the gene segments that, we posit, have been selected in germline evolution for their suppression of autoreactive potential.     

A novel approach to investigate tissue-specific trinucleotide repeat instability

Background

In Huntington's disease (HD), an expanded CAG repeat produces characteristic striatal neurodegeneration. Interestingly, the HD CAG repeat, whose length determines age at onset, undergoes tissue-specific somatic instability, predominant in the striatum, suggesting that tissue-specific CAG length changes could modify the disease process. Therefore, understanding the mechanisms underlying the tissue specificity of somatic instability may provide novel routes to therapies. However progress in this area has been hampered by the lack of sensitive high-throughput instability quantification methods and global approaches to identify the underlying factors.

Results

Here we describe a novel approach to gain insight into the factors responsible for the tissue specificity of somatic instability. Using accurate genetic knock-in mouse models of HD, we developed a reliable, high-throughput method to quantify tissue HD CAG repeat instability and integrated this with genome-wide bioinformatic approaches. Using tissue instability quantified in 16 tissues as a phenotype and tissue microarray gene expression as a predictor, we built a mathematical model and identified a gene expression signature that accurately predicted tissue instability. Using the predictive ability of this signature we found that somatic instability was not a consequence of pathogenesis. In support of this, genetic crosses with models of accelerated neuropathology failed to induce somatic instability. In addition, we searched for genes and pathways that correlated with tissue instability. We found that expression levels of DNA repair genes did not explain the tissue specificity of somatic instability. Instead, our data implicate other pathways, particularly cell cycle, metabolism and neurotransmitter pathways, acting in combination to generate tissue-specific patterns of instability.

Conclusion

Our study clearly demonstrates that multiple tissue factors reflect the level of somatic instability in different tissues. In addition, our quantitative, genome-wide approach is readily applicable to high-throughput assays and opens the door to widespread applications with the potential to accelerate the discovery of drugs that alter tissue instability.