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. 2023 Apr 8;24(8):6940.
doi: 10.3390/ijms24086940.

Function Analysis of Cholesterol 7-Desaturase in Ovarian Maturation and Molting in Macrobrachium nipponense: Providing Evidence for Reproductive Molting Progress

Jisheng Wang  1 , Sufei Jiang  1   2 , Wenyi Zhang  2 , Yiwei Xiong  2 , Shubo Jin  2 , Dan Cheng  1 , Yalu Zheng  1 , Hui Qiao  1   2 , Hongtuo Fu  1   2
Affiliations

Function Analysis of Cholesterol 7-Desaturase in Ovarian Maturation and Molting in Macrobrachium nipponense: Providing Evidence for Reproductive Molting Progress

Jisheng Wang et al. Int J Mol Sci. .

Abstract

The Cholesterol 7-desaturase gene plays an important role in insect ecdysone synthesis, but its role in ovarian development has not been reported. In this study, characteristics and the phylogenetic relationship of Cholesterol 7-desaturase were identified by bioinformatics. qPCR showed that the Mn-CH7D gene was highly expressed in the ovary, which was much higher than that in other tissues, and the expression level of Mn-CH7D reached the highest level at the third stage of the ovarian development stage (O-III). During embryonic development, the Mn-CH7D gene expression was highest in the zoea stage. The function of the Mn-CH7D gene was explored by RNA interference. The experimental group was injected with Mn-CH7D dsRNA through the pericardial cavity of M. nipponense, while the control group was injected with the same volume of dsGFP. Statistical analysis of gonadal development and GSI calculation showed that the silencing of Mn-CH7D resulted in the suppression of gonadal development. In addition, the molting frequency of the experimental group was significantly lower than that of the control group during the second molting cycle after silencing Mn-CH7D. On the seventh day after silencing, ecdysone content in the experimental group was significantly reduced. These results demonstrated that the Mn-CH7D gene played a dual role in ovarian maturation and molting of M. nipponense.

Keywords: Cholesterol 7-desaturase; Macrobrachium nipponense; molting; ovarian maturation.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Full cDNA sequence and predicted amino acid sequence of Mn-CH7D. The start codon ATG and the stop codon TAG are marked with red. The gray shades are two conserved domains, in order: the Rieske [2Fe-2S] domain and the non-heme iron binding domain. The black box represents the tailing signal and poly(A). The stop codon TAG in the amino acid sequence is represented by an asterisk (*).
Figure 2
Figure 2
Alignment of the deduced amino acid sequence of Mn-CH7D with other species. N-myristoylation sites are marked by red boxes, and Casein kinase II phosphorylation sites are marked by yellow boxes.
Figure 3
Figure 3
Phylogenetic tree connecting the Mn-CH7D amino acid sequence of Macrobrachium nipponense and other species. The numbers in brackets indicate the GenBank accession numbers. The numbers shown at the branches indicate the bootstrap values (%).
Figure 4
Figure 4
The expression pattern of the Mn-CH7D gene in different tissues (A), different stages of ovarian maturation (B), and developmental stages (C) of Macrobrachium nipponense were measured by qPCR. E: eyestalk, Br: cerebral ganglion, H: heart, He: hepatopancreas, G: gill, M: muscle, O: ovary; Data are presented as the mean ± SD (n = 6). Different letters indicate significant differences. p < 0.05 was considered to be statistically significant.
Figure 5
Figure 5
Localization of the expression of Mn-CH7D mRNA in Macrobrachium nipponense ovaries using in situ hybridization. OC: oocyte; N: nucleus; CM: cytoplasmic membrane; Y: yolk granule; FC: follicle cell; Scale bars: High magnification 400×.
Figure 6
Figure 6
Expression levels of Mn-CH7D in ovaries of Macrobrachium nipponense after injection with Mn-CH7D dsRNA. Data are shown as mean ± SD (n = 6). “**” indicates the significance of the differences (p < 0.01).
Figure 7
Figure 7
The percentage of development stages past stage III (A) and GSI (%) (B) of female Macrobrachium nipponense after injection with Mn-CH7D dsRNA. Data are shown as mean ± SD (n = 6). “*” indicates the significance of the differences (p < 0.05). “**” indicates the significance of the differences (p < 0.01).
Figure 8
Figure 8
The molting frequency (A) and ecdysone content (B) of Macrobrachium nipponense after injection with Mn-CH7D dsRNA. Data are shown as mean ± SD (n = 6). “**” indicates the significance of the differences (p < 0.01).
Figure 9
Figure 9
Histological observation of ovary of Macrobrachium nipponense after injection with Mn-CH7D dsRNA. OC: oocyte; N: nucleus; CM: cytoplasmic membrane; Scale bars: 400×.
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