Identification and circadian expression patterns of circadian clock gene <i>Tim</i> and its effects on sex hormone secretion in <i>Sinonovacula constricta</i>

Li Qijun; Zheng Xiangyang; Liu Yanzi; Yan Yuxiao; Chen Yanlei; Yao Hanhan; Dong Yinghui

doi:10.12284/hyxb2023181

Volume 45 Issue 12

Dec. 2023

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Li Qijun，Zheng Xiangyang，Liu Yanzi, et al. Identification and circadian expression patterns of circadian clock gene Tim and its effects on sex hormone secretion in Sinonovacula constricta[J]. Haiyang Xuebao，2023, 45(12)：101–111 doi: 10.12284/hyxb2023181

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Li Qijun，Zheng Xiangyang，Liu Yanzi, et al. Identification and circadian expression patterns of circadian clock gene Tim and its effects on sex hormone secretion in Sinonovacula constricta[J]. Haiyang Xuebao，2023, 45(12)：101–111 doi: 10.12284/hyxb2023181

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Identification and circadian expression patterns of circadian clock gene Tim and its effects on sex hormone secretion in Sinonovacula constricta

doi: 10.12284/hyxb2023181

1.
Zhejiang Key Laboratory of Aquatic Germplasm Resources, College of Biological & Environmental Sciences, Zhejiang Wanli University, Ningbo 315100, China
2.
Ninghai Marine Biological Seed Industry Research Institute, Ninghai 315604, China

Received Date: 2023-07-14
Rev Recd Date: 2023-11-27

Available Online: 2024-01-12

Publish Date: 2023-12-01

Abstract

Abstract

Tim (Timeless) is the second core circadian clock gene identified after the discovery of Per, which plays an important role in the regulation of animal reproduction. In order to explore the molecular mechanisms of Tim to regulate spawning in Sinonovacula constricta, the cDNA sequence of Tim was cloned and analyzed, the circadian expression patterns of Tim gene/protein in different tissues were analyzed by qRT-PCR and immunofluorescence technology, and the regulatory effects of Tim on sex hormone secretion were investigated by siRNA technology. The ORF sequence of Tim was 3 090 bp, encoding 1 029 amino acids and containing three conserved domains (TIMELESS superfamily、TIMELESS_C and FIil). Tim was expressed in test tissues, and showed higher expression levels in the hepatopancreas, siphon, ovary and testis. The expression levels of Tim were higher at 12:00–18:00 in the daytime and lower at 00:00–06:00 in the night time, which exhibited circadian rhythmic expression patterns in the ovary and testis. Tim protein was mainly expressed in hepatocytes of the hepatopancreas, epithelial cells of siphons, mature oocytes of the ovary, and spermatids of the testis. And protein fluorescence signals of Tim were slightly stronger at 12:00 during the day than at 00:00 during the night in the ovary and testis. After siRNA injection, the expression levels of Tim significantly decreased from the 3rd to 7th day in the siRNA treatment groups of the ovary and testis (P < 0.05). Meanwhile, the contents of testosterone in the testis and estradiol in the ovary decreased significantly from the 1st to 9th day after siRNA injection (P < 0.01). Consequently, it could be concluded that Tim gene/protein exhibited circadian rhythmic expression patterns in the ovary and testis, and the contents of sex hormones significantly decreased after Tim interference, which suggested that Tim might regulate the nocturnal spawning by mediating the secretion of sex hormones in S. constricta. These findings preliminarily explored the regulatory effects of circadian clock gene Tim on nocturnal spawning of S. constricta, which will provide important application value for in-depth analysis of spawning mechanism and optimizing industrialized seedling breeding technology in mollusks.
- Sinonovacula constricta,
- circadian clock,
- Tim,
- gene expression,
- siRNA,
- sex hormone

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References(34)

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