Project description:In poultry, in vitro derived primordial germ cells (PGCs) represent an important tool for management of genetic resources. However, several studies have highlighted sexual differences exhibited by PGCs through in vitro steps, which may compromise their reproductive capacities. To understand this phenomenon, we compared the proteome of pregonadal chicken male (ZZ) and female (ZW) PGCs expanded in vitro by quantitative proteomic analysis using a GeLC-MS/MS strategy. The proteins found to be differentially abundant in chicken male and female PGCs indicated their early sexual identity. Many of the proteins up-accumulated in male PGCs were encoded by genes strongly enriched in the sexual chromosome Z. This suggests that the known lack of dosage compensation of the transcription of Z-linked genes between sexes persists at protein level in PGCs, and that this may be a key factor of their autonomous sex differentiation. Male and female PGCs up-accumulated protein sets were associated with differential biological processes, and contained proteins biologically relevant for male and female germ cell development respectively. This study presents first evidence on early predetermined sex specific cell fate of chicken PGCs that will help to understand their sexual physiological specificities and enable more precise sex-specific adaptation of in vitro culture conditions.
Project description:Chicken primordial germ cells (PGCs) have an epigenetic signature which differs from the one that mammalian PGCs acquire with their epigenome reprogramming during early embryonic development. In particular, chicken PGCs display a high global amount of histone H3 lysine 9 trimethylation (H3K9me3) compared to somatic cell types. We performed the genome-wide profiling of H3K9me3 and the transcriptome analysis on chicken PGCs compared to embryonic stem cells (ESCs) as a closely related, non germinal cell type.
Project description:Background: Genes, RNAs, and proteins play important roles during germline development. However, the functions of non-coding RNAs (ncRNAs) on germline development remain unclear in avian species. Recent high-throughput techniques have identified several classes of ncRNAs, including micro RNAs (miRNAs), small-interfering RNAs (siRNAs), and PIWI-interacting RNAs (piRNAs). These ncRNAs are functionally important in the genome, however, the identification and annotation of ncRNAs in a genome is challenging. The aim of this study was to identify different types of small ncRNAs particularly piRNAs, and the role of piRNA pathway genes in the protection of chicken primordial germ cells (PGCs). Results: At first, we performed next-generation sequencing to identify ncRNAs in chicken PGCs, and we performed ab initio predictive analysis to identify putative piRNAs in PGCs. Then, we examined the expression of three repetitive sequence-linked piRNAs and 14 genic-transcript-linked piRNAs along with their linked genes using real-time PCR. All piRNAs and their linked genes were highly expressed in PGCs. Subsequently, we knocked down two known piRNA pathway genes of chicken, PIWI-like protein 1 (CIWI) and 2 (CILI), in PGCs using siRNAs. After knockdown of CIWI and CILI, we examined their effects on the expression of six putative piRNA-linked genes and DNA double-strand breakage in PGCs. The knockdown of CIWI and CILI upregulated chicken repetitive 1 (CR1) element and RAP2B, a member of RAS oncogene family, and increased DNA double-strand breakage in PGCs. Conclusions: Our results increase the understanding of PGC-expressed ncRNAs and the role of piRNA pathway genes in the protection of germ cells.
Project description:The existence of conventional dendritic cells (cDCs) has not yet been demonstrated outside mammals. In this paper, we identified bona fide cDCs in chicken spleen. Comparative profiling of global and of immune response gene expression, morphology, and T cell activation properties show that cDCs and macrophages (MPs) exist as distinct mononuclear phagocytes in chicken, resembling their human and mouse cell counterparts. Using computational analysis, core gene expression signatures for cDCs, MPs, T and B cells across chicken, human and mouse were established, which will facilitate the identification of these subsets in other vertebrates. Overall this study, by extending the newly uncovered cDC and MP paradigm to chicken, suggests that the generation of these two phagocyte lineages occurred before the reptile to mammal and bird transition in evolution. It opens avenues for the design of new vaccines and neutraceuticals that are mandatory for the sustained supply of poultry products in the expanding human population.