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The germline-enriched Ppp1r36 promotes autophagy.

ABSTRACT: Spermatogenesis is a highly regulated process during which haploid sperm cells are generated. Although autophagy is involved in the spermatogenesis process, the molecular pathways and regulations of autophagy in germ cell development remain elusive. Here, we showed that Ppp1r36, a regulatory subunit of protein phosphatase 1, is expressed during gonadal development, mainly in testes during spermatogenesis. Autophagy protein LC3 (microtubule associated protein 1 light chain 3), especially its active form LC3-II, had a similar expression pattern to Ppp1r36. Moreover, LC3-II level and puncta analysis showed that autophagy is up-regulated around 21?dpp (day postpartum) in postnatal testis, indicating a potential role of autophagy during the first wave of spermatogenesis. We demonstrated that Ppp1r36 promotes autophagosome formation upon starvation induction. Further autophagy flux analysis using a tandem fluorescent indicator, mCherry-GFP-LC3, confirmed that Ppp1r36 participated in autophagy. We further determined that Ppp1r36 is associated with Atg16L1 (autophagy related 16-like 1) in autophagy of starvation induction. Thus, our results uncover a potential role of the regulatory subunit Ppp1r36 of protein phosphatase 1 in enhancing autophagy during spermatogenesis.

SUBMITTER: Zhang Q

PROVIDER: S-EPMC4838909 | biostudies-other | 2016 Apr

REPOSITORIES: biostudies-other

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The germline-enriched Ppp1r36 promotes autophagy.

Zhang Qinghua Q Gao Maomao M Zhang Ying Y Song Ying Y Cheng Hanhua H Zhou Rongjia R

Scientific reports 20160421

Spermatogenesis is a highly regulated process during which haploid sperm cells are generated. Although autophagy is involved in the spermatogenesis process, the molecular pathways and regulations of autophagy in germ cell development remain elusive. Here, we showed that Ppp1r36, a regulatory subunit of protein phosphatase 1, is expressed during gonadal development, mainly in testes during spermatogenesis. Autophagy protein LC3 (microtubule associated protein 1 light chain 3), especially its acti ...[more]

PMID: 27098880

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Project description:The present study aims to evaluate season- and reproductive-stage dependent variation in culture characteristics and expression of pluripotency and adhesion markers in caprine-male germline stem cells (cmGSCs). For this, testes from pre-pubertal (4-6 months) and adult (~ 2 years) bucks during non-breeding (July-August; n = 4 each) and breeding (October-November; n = 4 each) seasons were used to isolated testicular cells by two-step enzymatic digestion. After cmGSCs enrichment by multiple methods (differential platting, Percoll density gradient centrifugation, and MACS), cell viability of CD90+ cells was assessed before co-cultured onto the Sertoli cell feeder layer up to 3rd-passage (P-3). The culture characteristics of cmGSCs were compared during primary culture (P-0) and P-3 with different assays [BrdU-assay (proliferation), MTT-assay (senescence), and Cluster-forming activity-assay] and transcript expression analyses by qRT-PCR. Moreover, the co-localization of UCHL-1, CD90, and DBA was examined by a double-immunofluorescence method. In adult bucks, significantly (p < 0.05) higher cell numbers with the ability to proliferate faster and form a greater number of cell clusters, besides up-regulation of pluripotency and adhesion markers expression were observed during the breeding season than the non-breeding season. In contrast, such season-dependent variation was lacking in pre-pubertal bucks. The expression of transcripts during non-breeding seasons was significantly (p < 0.05) higher in pre-pubertal cmGSCs than in adult cells (UCHL-1 = 2.38-folds; CD-90 = 6.66-folds; PLZF = 20.87-folds; ID-4 = 4.75-folds; E-cadherin = 3.89-folds and β1-integrin = 5.70-folds). Overall, the reproductive stage and season affect the population, culture characteristics, and expression of pluripotency and adhesion specific markers in buck testis. These results provide an insight to develop an efficient system for successful cell culture processes targeting cmGSCs.Supplementary informationThe online version contains supplementary material available at 10.1007/s10616-021-00515-x.

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The germline-enriched Ppp1r36 promotes autophagy.

Publications

The germline-enriched Ppp1r36 promotes autophagy.

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