CFTR is required for the migration of primordial germ cells during zebrafish early embryogenesis

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene affect fertility in both sexes. However, the involvement of CFTR in regulating germ cell development remains largely unknown. Here, we used zebrafish model to investigate the role of CFTR in primordial germ cells (PGCs) development. We generated a cftr frameshift mutant zebrafish line using CRISPR/Cas9 technique and investigated the migration of PGCs during early embryo development. Our results showed that loss of Cftr impairs the migration of PGCs from dome stages onward. The migration of PGCs was also perturbed by treatment of CFTRinh-172, a gating-speciﬁc CFTR channel inhibitor. Moreover, defected PGCs migration in cftr mutant embryos can be partially rescued by injection of WT but not other channel-defective mutant cftr mRNAs. Finally, we observed the elevation of cxcr4b, cxcl12a, rgs14a and ca15b, key factors involved in zebrafish PGCs migration, in cftr-mutant zebrafish embryos. Taken together, the present study revealed an important role of CFTR acting as an ion channel in regulating PGCs migration during early embryogenesis. Defect of which may impair germ cell development through elevation of key factors involved in cell motility and response to chemotactic gradient in PGCs.

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Disruption of Tcfap2c in Primordial Germ Cells using Prdm1-Cre Prevents Germ Cell Development

Biology of Reproduction ◽

10.1093/biolreprod/78.s1.64a ◽

2008 ◽

Vol 78 (Suppl_1) ◽

pp. 64-64

Author(s):

Jillian Guttormsen ◽

Gerrit J. Bouma ◽

Frances Bhushan ◽

Trevor Williams ◽

Quinton A. Winger

Keyword(s):

Germ Cell ◽

Germ Cells ◽

Primordial Germ Cells ◽

Cell Development ◽

Germ Cell Development

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Isolation of the murine cyclin B2 cDNA and characterization of the lineage and temporal specificity of expression of the B1 and B2 cyclins during oogenesis, spermatogenesis and early embryogenesis

Development ◽

10.1242/dev.118.1.229 ◽

1993 ◽

Vol 118 (1) ◽

pp. 229-240 ◽

Cited By ~ 1

Author(s):

D.L. Chapman ◽

D.J. Wolgemuth

Keyword(s):

Germ Cell ◽

Germ Cells ◽

Northern Blot ◽

Inner Cell Mass ◽

Cell Mass ◽

Cell Development ◽

Early Embryogenesis ◽

Northern Blot Hybridization ◽

Germ Cell Development

A cDNA encoding the murine cyclin B2 (cycB2) was isolated from an adult mouse testis cDNA library as part of studies designed to identify cyclins involved in murine germ cell development. This cycB2 cDNA was then used to examine the pattern of cycB2 expression during male and female germ cell development and in early embryogenesis, and to compare this expression with the previously characterized expression of cycB1. A single 1.7 kb cycB2 transcript was detected by northern blot hybridization analysis of total RNA isolated from midgestation embryos and various adult tissues. Northern blot and in situ hybridization analyses revealed that cycB2 expression in the testis was most abundant in the germ cells, specifically in pachytene spermatocytes. This is in contrast to the highest levels of expression of cycB1 being present in early spermatids. In situ analysis of the ovary revealed cycB2 transcripts in both germ cells and somatic cells, specifically in the oocytes and granulosa cells of growing and mature follicles. The pattern of cycB1 and cycB2 expression in ovulated and fertilized eggs was also examined. While the steady state level of cycB1 and cycB2 signal remained constant in oocytes and ovulated eggs, signal of both appeared to decrease following fertilization. In addition, both cycB1 and cycB2 transcripts were detected in the cells of the inner cell mass and the trophectoderm of the blastocyst. These results demonstrate that there are lineage- and developmental-specific differences in the pattern of the B cyclins in mammalian germ cells, in contrast to the co-expression of B cyclins in the early conceptus.

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Single cell RNA sequencing reveals the landscape of early female germ cell development

10.1101/2020.05.09.085845 ◽

2020 ◽

Cited By ~ 2

Author(s):

Zheng-Hui Zhao ◽

Jun-Yu Ma ◽

Tie-Gang Meng ◽

Zhen-Bo Wang ◽

Wei Yue ◽

...

Keyword(s):

Germ Cell ◽

Germ Cells ◽

Primordial Germ Cells ◽

Cell Development ◽

Developmental Trajectory ◽

Special Focus ◽

Germ Cell Development ◽

New Information ◽

Resolution Level ◽

Female Germ Cell

ABSTRACTFemale germ cell development consists of complex events including sex determination, meiosis initiation, retardation and resumption. During early oogenesis, the asynchrony of the transition from mitosis to meiosis results in heterogeneity in the female germ cell populations at a certain embryonic stage, which limits the studies of meiosis initiation and progression at a higher resolution level. Here, we investigated the transcriptional profiles of 19363 single germ cells collected from E12.5, E14.5 and E16.5 mouse fetal ovaries. Clustering analysis identified seven groups and defined dozens of corresponding transcription factors, providing a global view of cellular differentiation from primordial germ cells towards meiocytes. Further, we explored the dynamics of gene expression within the developmental trajectory with special focus on the mechanisms underlying meiotic initiation. We found that Dpy30 may be involved in the regulation of meiosis initiation at the epigenetic level. Our data provide key insights into the transcriptome features of peri-meiotic female germ cells, which offers new information not only on meiosis initiation and progression but also on screening pathogenic mutations in meiosis-associated diseases.

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301 IMPRINTED GENE EXPRESSION PATTERNS IN CULTURED PRIMORDIAL GERM CELLS DERIVED FROM PORCINE EMBRYOS BETWEEN DAY 25 AND 30

Reproduction Fertility and Development ◽

10.1071/rdv23n1ab301 ◽

2011 ◽

Vol 23 (1) ◽

pp. 247

Author(s):

C. K. Lee ◽

C. H. Park ◽

K. J. Uh ◽

J. K. Park ◽

H. S. Kim

Keyword(s):

Gene Expression ◽

Germ Cell ◽

Germ Cells ◽

Expression Patterns ◽

Primordial Germ Cells ◽

Cell Development ◽

Epigenetic Reprogramming ◽

Expression Level ◽

Imprinted Gene ◽

Germ Cell Development

Although epigenetic reprogramming during germ cell development has extensively been studied in the mouse, little is known about the timing of imprinting accomplishment in the pig. Therefore, the aim of this study was to investigate the timing of epigenetic reprogramming in porcine primordial germ cells (PGC) through short-term culture as well as to confirm the differences of epigenetic features between PGC and embryonic germ cells (EGC). The present study aimed to investigate mRNA expression pattern of imprinted genes in cultured PGC that were derived from embryos at Day 25 to 30 of pregnancy. The overall expression pattern can be separated into 3 different features: no changes in all PGC regardless of embryonic day, gradual increases through embryonic day, and extreme increases in PGC derived from embryos at Day 30. Of 8 imprinted genes, Grb10, Peg10, and Snrpn transcripts were retained through all stages. The expression level of H19, Igf2, Igf2r, and Peg1/Mest progressively increased in PGC derived at later stages of embryo development. Interestingly, Xist and Nnat transcripts showed extremely high levels of expression in PGC derived from embryo at Day 30 of pregnancy. The expression level of these transcripts was 3- to 4-fold higher in PGC derived from embryo at Day 30 of pregnancy compared with that of the others. Our results indicated that analysis of the imprinted gene expression level in short-term cultured PGC may in part reflect epigenetic modifications during germ cell development in pig. This study could help us understand how epigenetic reprogramming is maintained or lost during germ cell development. Further study will be carried out to compare imprinted gene expression patterns between the porcine PGC and EGC. This work was supported by the BioGreen 21 Program (#20070401034031, #20080401034031), Rural Development Administration, Republic of Korea (HK).

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Tissue non-specific alkaline phosphatase is expressed in both embryonic and extraembryonic lineages during mouse embryogenesis but is not required for migration of primordial germ cells

Development ◽

10.1242/dev.121.5.1487 ◽

1995 ◽

Vol 121 (5) ◽

pp. 1487-1496 ◽

Cited By ~ 12

Author(s):

G.R. MacGregor ◽

B.P. Zambrowicz ◽

P. Soriano

Keyword(s):

Alkaline Phosphatase ◽

Germ Cell ◽

Germ Cells ◽

Primordial Germ Cells ◽

Primordial Germ Cell ◽

Embryonic Stem ◽

Blastocyst Stage ◽

Early Embryo ◽

Specific Alkaline Phosphatase

Mouse primordial germ cells express tissue non-specific alkaline phosphatase (TNAP) during development, but the widespread expression of another alkaline phosphatase gene in the early embryo limits the potential use of this marker to trace germ cells. To attempt to identify germ cells at all stages during embryonic development and to understand the role of TNAP in germ cell ontogeny, mice carrying a beta geo (lacZ/neor) disrupted allele of the TNAP gene were generated by homologous recombination in embryonic stem cells. Using beta-galactosidase activity, the embryonic pattern of TNAP expression was examined from the blastocyst stage to embryonic day 14. Results indicate that primordial germ cell progenitors do not express TNAP prior to gastrulation although at earlier times TNAP expression is found in an extraembryonic lineage destined to form the chorion. In homozygous mutants, primordial germ cells appear unaffected indicating that TNAP is not essential for their development or migration.

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hnRNPH1 recruits PTBP2 and SRSF3 to cooperatively modulate alternative pre-mRNA splicing in germ cells and is essential for spermatogenesis and oogenesis

10.21203/rs.3.rs-1060705/v1 ◽

2021 ◽

Author(s):

Shuiqiao Yuan ◽

Shenglei Feng ◽

Jinmei Li ◽

Hui Wen ◽

Kuan Liu ◽

...

Keyword(s):

Alternative Splicing ◽

Germ Cell ◽

Germ Cells ◽

Rna Binding ◽

Cell Development ◽

Mrna Splicing ◽

Conditional Knockout ◽

Cell Junction ◽

Germ Cell Development ◽

Splicing Regulators

Abstract Coordinated regulation of alternative pre-mRNA splicing is essential for germ cell development. However, the molecular mechanism underlying that control alternative mRNA expression during germ cell development remains poorly understood. Herein, we showed that hnRNPH1, an RNA-binding protein, is highly expressed in the reproductive system and localized in the chromosomes of meiotic cells but excluded from the XY body in pachytene spermatocytes and recruits the splicing regulators PTBP2 and SRSF3 and cooperatively regulates the alternative splicing of the critical genes that are required for spermatogenesis. Conditional knockout Hnrnph1 in spermatogenic cells caused many abnormal splicing events that affect genes related to meiosis and communication between germ cells and Sertoli cells, characterized by asynapsis of chromosomes and impairment of germ-Sertoli communications, ultimately leading to male sterility. We further showed that hnRNPH1 could directly bind to SPO11 and recruit the splicing regulators PTBP2 and SRSF3 to regulate the alternative splicing of the target genes cooperatively. Strikingly, Hnrnph1 germline-specific mutant female mice were also infertile, and Hnrnph1-deficient oocytes exhibited a similar defective synapsis and cell-cell junction as shown in Hnrnph1-deficient male germ cells. Collectively, our data reveal an essential role for hnRNPH1 in regulating pre-mRNA splicing during spermatogenesis and oogenesis and support a molecular model whereby hnRNPH1 governs a network of alternative splicing events in germ cells via recruiting PTBP2 and SRSF3.

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Ovol2, a zinc finger transcription factor, is dispensable for spermatogenesis in mice

Reproductive Biology and Endocrinology ◽

10.1186/s12958-019-0542-3 ◽

2019 ◽

Vol 17 (1) ◽

Cited By ~ 1

Author(s):

Jin Zhang ◽

Juan Dong ◽

Weibing Qin ◽

Congcong Cao ◽

Yujiao Wen ◽

...

Keyword(s):

Transcription Factor ◽

Germ Cell ◽

Zinc Finger ◽

Germ Cells ◽

Cell Development ◽

Knockout Mouse Model ◽

Zinc Finger Transcription Factor ◽

Germ Cell Development ◽

Embryonic Germ Cells ◽

Protein Levels

AbstractOvol2, a mouse homolog of Drosophila ovo, was identified as a zinc finger transcription factor predominantly expressed in testis. However, the function of Ovol2 in postnatal male germ cell development remains enigmatic. Here, we firstly examined the mRNA and protein levels of Ovol2 in developing mouse testes by RT-qPCR and western blot and found that both mRNA and protein of Ovol2 are continually expressed in postnatal developing testes from postnatal day 0 (P0) testes to adult testes (P56) and exhibits its higher level at adult testis. Further testicular immuno-staining revealed that OVOL2 is highly expressed in the spermatogonia, spermatocytes and round spermatids. Interestingly, our conditional ovol2 knockout mouse model show that loss of ovol2 in embryonic germ cells does not affect fecundity in mice. Our data also show that Ovol1 may have compensated for the loss of Ovol2 functions in germ cells. Overall, our data indicate that ovol2 is dispensable for germ cell development and spermatogenesis.

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Proteomic and metabolomic analyses uncover sex-specific regulatory pathways in mouse fetal germline differentiation†

Biology of Reproduction ◽

10.1093/biolre/ioaa115 ◽

2020 ◽

Vol 103 (4) ◽

pp. 717-735

Author(s):

Yohei Hayashi ◽

Masaru Mori ◽

Kaori Igarashi ◽

Keiko Tanaka ◽

Asuka Takehara ◽

...

Keyword(s):

Oxidative Phosphorylation ◽

Germ Cell ◽

Germ Cells ◽

Cell Development ◽

Nucleotide Synthesis ◽

Male Germ Cells ◽

Regulatory Pathways ◽

Germ Cell Development ◽

Germ Cell Differentiation ◽

Germline Differentiation

Abstract Regulatory mechanisms of germline differentiation have generally been explained via the function of signaling pathways, transcription factors, and epigenetic regulation; however, little is known regarding proteomic and metabolomic regulation and their contribution to germ cell development. Here, we conducted integrated proteomic and metabolomic analyses of fetal germ cells in mice on embryonic day (E)13.5 and E18.5 and demonstrate sex- and developmental stage-dependent changes in these processes. In male germ cells, RNA processing, translation, oxidative phosphorylation, and nucleotide synthesis are dominant in E13.5 and then decline until E18.5, which corresponds to the prolonged cell division and more enhanced hyper-transcription/translation in male primordial germ cells and their subsequent repression. Tricarboxylic acid cycle and one-carbon pathway are consistently upregulated in fetal male germ cells, suggesting their involvement in epigenetic changes preceding in males. Increased protein stability and oxidative phosphorylation during female germ cell differentiation suggests an upregulation of aerobic energy metabolism, which likely contributes to the proteostasis required for oocyte maturation in subsequent stages. The features elucidated in this study shed light on the unrevealed mechanisms of germ cell development.

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