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Updated: Jul 26, 2025

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Published on: November 29, 2016
mTORC1/rpS6 and mTORC2/PKC regulate spermatogenesis through Arp3-mediated actin microfilament organization in
Zhen-Fang Li1, Hong-Yu Qi1, Jia-Ming Wang1
1The Sperm Laboratory, College of Life Sciences, Zhejiang University, Hangzhou, 310058, China.
Abstract:
Mammalian target of rapamycin (mTOR) is a crucial signaling protein regulating a range of cellular events. Numerous studies have reported that the mTOR pathway is related to spermatogenesis in mammals. However, its functions and underlying mechanisms in crustaceans remain largely unknown. mTOR exists as two multimeric functional complexes termed mTOR complex 1 (mTORC1) and mTORC2. Herein, we first cloned ribosomal protein S6 (rpS6, a downstream molecule of mTORC1) and protein kinase C (PKC, a downstream effector of mTORC2) from the testis of Eriocheir sinensis. The dynamic localization of rpS6 and PKC suggested that both proteins may be essential for spermatogenesis. rpS6/PKC knockdown and Torin1 treatment led to defects in spermatogenesis, including germ cell loss, retention of mature sperm and empty lumen formation. In addition, the integrity of the testis barrier (similar to the blood-testis barrier in mammals) was disrupted in the rpS6/PKC knockdown and Torin1 treatment groups, accompanied by changing in expression and distribution of junction proteins. Further study demonstrated that these findings may result from the disorganization of filamentous actin (F-actin) networks, which were mediated by the expression of actin-related protein 3 (Arp3) rather than epidermal growth factor receptor pathway substrate 8 (Eps8). In summary, our study illustrated that mTORC1/rpS6 and mTORC2/PKC regulated spermatogenesis via Arp3-mediated actin microfilament organization in E. sinensis.
Insights
The mammalian target of rapamycin (mTOR) pathway regulates spermatogenesis in crustaceans. This study shows mTORC1/rpS6 and mTORC2/PKC signaling influence sperm development and testis barrier integrity via actin organization.
Area of Science:
- Reproductive Biology
- Cell Signaling
- Crustacean Research
Background:
- The mammalian target of rapamycin (mTOR) pathway is vital for cellular functions, including mammalian spermatogenesis.
- Its role in crustacean reproduction is largely unexplored.
- mTOR functions via two complexes: mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2).
Purpose of the Study:
- To investigate the function of mTORC1 and mTORC2 signaling in spermatogenesis of the crab Eriocheir sinensis.
- To elucidate the underlying molecular mechanisms, particularly concerning testis barrier integrity and actin organization.
Main Methods:
- Cloning of downstream effectors: ribosomal protein S6 (rpS6) for mTORC1 and protein kinase C (PKC) for mTORC2.
- Functional analysis using knockdown of rpS6/PKC and treatment with the mTOR inhibitor Torin1.
- Assessment of spermatogenesis, testis barrier integrity, junction proteins, and filamentous actin (F-actin) organization.
Main Results:
- Knockdown of rpS6/PKC or Torin1 treatment caused spermatogenesis defects, including germ cell loss and sperm retention.
- Testis barrier integrity was disrupted, with altered expression of junction proteins.
- These disruptions were linked to the disorganization of F-actin networks, mediated by actin-related protein 3 (Arp3).
Conclusions:
- The mTORC1/rpS6 and mTORC2/PKC pathways are essential for spermatogenesis in E. sinensis.
- These pathways regulate sperm development and testis barrier function through Arp3-mediated actin organization.
- This study reveals novel insights into mTOR signaling in invertebrate reproduction.
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