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Simple Summary: Neudesin neurotrophic factor (NENF) is a secreted protein that was significantly inhibited in the fat-muscle co-culture system in our previous study. However, studies on NENF regulation of bovine muscle development and involvement in the cross-talk between adipose tissue and skeletal muscle have not been reported. Hence, the aim of this study was to clarify the roles of NENF in bovine myoblast and preadipocyte differentiation. In this study, we first examined the spatial expression patterns of NENF in different tissues and found that NENF was highly expressed in the muscle of four-day-old and 24-month-old Qinchuan cattle. Compared with 4-day-old Qinchuan cattle, the expression level of NENF was significantly up-regulated in 24-month-old bovine adipose tissue. Then, we detected the expression pattern of the NENF gene in bovine preadipocyte and myoblast differentiation and found that the expression of NENF mRNA peaks at day 6 during preadipocyte differentiation and peaks at day 4 during myoblast differentiation. Furthermore, we found that the endogenous knockdown of NENF inhibited the differentiation of preadipocytes and promoted the differentiation of myoblasts. These findings not only lay the foundation for the construction of regulatory pathways during fat and muscle differentiation but also provide a theoretical basis for molecular breeding of beef cattle. Neudesin neurotrophic factor (NENF) is a secreted protein that is essential in multiple biological processes, including neural functions, adipogenesis, and tumorigenesis. In our previous study, NENF was significantly inhibited in the bovine adipocytes-myoblasts co-culture system. However, studies on NENF regulation of bovine muscle development and involvement in the cross-talk between adipose tissue and skeletal muscle have not been reported. Hence, the aim of this study was to clarify the functional roles of NENF in bovine preadipocytes and myoblasts. Real-time quantitative PCR (RT-qPCR) was performed to examine the spatial expression patterns of NENF in different tissues, and the results showed that NENF was highly expressed in the muscle of four-day-old and 24-month-old Qinchuan cattle. Compared with four-day-old Qinchuan cattle, the expression level of NENF was significantly up-regulated in 24-month-old bovine adipose tissue. To explore the roles of NENF in bovine myoblast and preadipocyte differentiation, small interfering RNA (siRNA) targeting the NENF gene were transfected into bovine preadipocytes and myoblasts to knock down the expression of the NENF gene. The results showed that the knockdown of NENF in differentiating adipocytes attenuated lipid accumulation, decreased the mRNA expression of adipogenic key marker genes PPARγ, CEBPα, CEBPβ, FASN, and SCD1, and decreased the protein expression of PPARγ, CEBPα, and FASN. The formation of myotubes was significantly accelerated, and the mRNA expression levels of myogenic marker genes MYOD1, MYF5, MYF6, MEF2A, MEF2C, and CKM, and the protein expression levels of MYOD1, MYF6, MEF2A, and CKM were up-regulated in myoblasts where NENF was knocked down. In short, the knockdown of NENF inhibited preadipocyte differentiation and promoted myoblast myogenesis. [ABSTRACT FROM AUTHOR]

Simple Summary: In this article explore the role of the bovine TP53INP2 gene in adipocyte differentiation and its function in autophagy during the early stage of adipocyte differentiation. In our work we found that a novel, important autophagy related protein TP53INP2 can activate autophagy during the early stage of differentiation in bovine adipocytes and positively regulate adipocyte differentiation by affecting autophagy. Furthermore, we demonstrated that peroxisome proliferator-activated receptor gamma (PPARγ) also contributed to the function of TP53INP2 in modulating adipocyte differentiation. The study of the function of bovine TP53INP2 gene on adipocyte differentiation has not been reported, therefore, we have decided to focus on Qinchuan cattle, one of the five important cattle breeds in China. We propose that the TP53INP2 gene may affect the meat quality of Qinchuan cattle by regulating lipid deposition, and may shed new light on the developmental mechanisms of adipose development. Tumor protein p53 inducible nuclear protein 2 (TP53INP2) is a key positive regulator of autophagy, and it has been shown to modulate adipocyte differentiation. However, the molecular mechanism involved in autophagy regulation during adipocyte differentiation has not been clarified. Our experiments were intended to investigate whether TP53INP2 is involved in the regulation of autophagy during bovine adipocyte differentiation and how TP53INP2 affects the differentiation of bovine adipocytes. In our research, using RT-qPCR and Western blot methods, we found that the overexpression of TP53INP2 resulted in the upregulation of adipogenesis and autophagy-related genes, and autophagy flux and the degree of differentiation were detected by LipidTOX™ Deep Red Neutral Lipid staining and dansylcadaverine staining, respectively. The knockdown of TP53INP2 produced results that were the inverse of those produced by the overexpression of TP53INP2. Overall, our results suggested that TP53INP2 can activate autophagy during the early stage of differentiation in bovine adipocytes and positively regulate adipocyte differentiation by affecting autophagy. Additionally, peroxisome proliferator-activated receptor gamma (PPARγ) also contributed to the function of TP53INP2 in modulating adipocyte differentiation. [ABSTRACT FROM AUTHOR]