The journey of a generation: advances and promises in the study of primordial germ cell migration.

The germline provides the genetic and non-genetic information that passes from one generation to the next. Given this important role in species propagation, egg and sperm precursors, called primordial germ cells (PGCs), are one of the first cell types specified during embryogenesis. In fact, PGCs form well before the bipotential somatic gonad is specified. This common feature of germline development necessitates that PGCs migrate through many tissues to reach the somatic gonad. During their journey, PGCs must respond to select environmental cues while ignoring others in a dynamically developing embryo. The complex multi-tissue, combinatorial nature of PGC migration is an excellent model for understanding how cells navigate complex environments in vivo. Here, we discuss recent findings on the migratory path, the somatic cells that shepherd PGCs, the guidance cues somatic cells provide, and the PGC response to these cues to reach the gonad and establish the germline pool for future generations. We end by discussing the fate of wayward PGCs that fail to reach the gonad in diverse species. Collectively, this field is poised to yield important insights into emerging reproductive technologies.

Isolation of Undifferentiated Spermatogonia from Adult and Developing Mouse Testes.

In the mammalian testis, the mitotic complements of spermatogenic cells are spermatogonia, including spermatogonial stem cells (SSCs) which form the basis of life-long spermatogenesis and male fertility. Thus, investigating spermatogonia and subdivisions thereof is essential to increase our understanding of male germline development and infertility. This protocol describes the isolation of spermatogonia from both adult and developing [postnatal day 6 (P6)] mouse testes. Cell suspensions of the adult mouse testis from the Id4-Egfp transgenic mouse line are obtained through a two-step enzymatic digestion and are subjected to Percoll pre-enrichment before spermatogonia are isolated by selecting testis cells that are CD9bright and ID4-EGFP+ through FACS. For P6 mice, the testis is digested using trypsin-DNase, and spermatogonia are isolated by FACS selection of ID4-EGFP+ testis cells. In both cases, nearly pure populations of undifferentiated spermatogonia are obtained that can be further subdivided using additional parameters (e.g., EGFP intensity, cell surface protein immunostaining), and recovered for use in various downstream applications, such as biochemical analyses (e.g., transcriptome/epigenome), functional analyses by SSC transplantation or propagation in vitro.

Hallmarks of glycogene expression and glycosylation pathways in squamous and adenocarcinoma cervical cancer.

BACKGROUND: Dysregulation of glycogene expression in cancer can lead to aberrant glycan expression, which can promote tumorigenesis. Cervical cancer (CC) displays an increased expression of glycogenes involved in sialylation and sialylated glycans. Here, we show a comprehensive analysis of glycogene expression in CC to identify glycogene expression signatures and the possible glycosylation pathways altered. METHODS: First, we performed a microarray expression assay to compare glycogene expression changes between normal and cervical cancer tissues. Second, we used 401 glycogenes to analyze glycogene expression in adenocarcinoma and squamous carcinoma from RNA-seq data at the cBioPortal for Cancer Genomics. RESULTS: The analysis of the microarray expression assay indicated that CC displayed an increase in glycogenes related to GPI-anchored biosynthesis and a decrease in genes associated with chondroitin and dermatan sulfate with respect to normal tissue. Also, the glycogene analysis of CC samples by the RNA-seq showed that the glycogenes involved in the chondroitin and dermatan sulfate pathway were downregulated. Interestingly the adenocarcinoma tumors displayed a unique glycogene expression signature compared to squamous cancer that shows heterogeneous glycogene expression divided into six types. Squamous carcinoma type 5 (SCC-5) showed increased expression of genes implicated in keratan and heparan sulfate synthesis, glycosaminoglycan degradation, ganglio, and globo glycosphingolipid synthesis was related to poorly differentiated tumors and poor survival. Squamous carcinoma type 6 (SCC-6) displayed an increased expression of genes involved in chondroitin/dermatan sulfate synthesis and lacto and neolacto glycosphingolipid synthesis and was associated with nonkeratinizing squamous cancer and good survival. In summary, our study showed that CC tumors are not a uniform entity, and their glycome signatures could be related to different clinicopathological characteristics.

The Mammalian Spermatogenesis Single-Cell Transcriptome, from Spermatogonial Stem Cells to Spermatids.

Spermatogenesis is a complex and dynamic cellular differentiation process critical to male reproduction and sustained by spermatogonial stem cells (SSCs). Although patterns of gene expression have been described for aggregates of certain spermatogenic cell types, the full continuum of gene expression patterns underlying ongoing spermatogenesis in steady state was previously unclear. Here, we catalog single-cell transcriptomes for >62,000 individual spermatogenic cells from immature (postnatal day 6) and adult male mice and adult men. This allowed us to resolve SSC and progenitor spermatogonia, elucidate the full range of gene expression changes during male meiosis and spermiogenesis, and derive unique gene expression signatures for multiple mouse and human spermatogenic cell types and/or subtypes. These transcriptome datasets provide an information-rich resource for studies of SSCs, male meiosis, testicular cancer, male infertility, or contraceptive development, as well as a gene expression roadmap to be emulated in efforts to achieve spermatogenesis in vitro.

Expression analysis of ST3GAL4 transcripts in cervical cancer cells.

ST3GAL4 gene expression is altered in different cancer types, including cervical cancer. Several mRNA transcripts have been reported for this gene; however, their expression levels in cervical cancer have not been analyzed. ST3GAL4 encodes for ß­galactosidase α­2,3­sialyltransferase 4, involved in the biosynthesis of the tumour antigens sLe(x) and sulfo­sLe(x). The present study evaluated the presence of three mRNA variants (V1, V2 and V3) in cervical cancer cell lines, detecting the three variants. Additionally, the expression level of the V1 transcript of the ST3GAL4 gene was determined by reverse transcription­quantitative polymerase chain reaction in cervical cell lines and in normal, premalignant and cervical cancer tissue. The V1 transcript of the ST3GAL4 demonstrated significant decreased expression in premalignant and malignant cervical tissues. The results suggested that deregulation of this gene could occur prior to the presence of cancer and demonstrated the importance of evaluating the expression level of V1, and its association with disease progression.