Retinoblastoma 1 (RB1) modulates the proliferation of chicken preadipocytes

Abstract

18Retinoblastoma 1 (RB1) has been extensively studied in mammalian species, but its 19 function in avian species is unclear. The objective of this study was to reveal the role of 20 chicken RB1 (Gallus gallus RB1, gRB1) in the proliferation of preadipocytes. In the current 21 study, quantitative real-time PCR analysis showed that the expression levels of gRB1 22 transiently increased during the proliferation of preadipocytes. The MTT assay showed that 23 gRB1 overexpression suppressed preadipocyte proliferation, and gRB1 interference promoted 24 preadipocyte proliferation. Additionally, cell-cycle analysis indicated that gRB1 may play a 25 crucial role in the G1/S transition. Consistently, gene expression analysis showed that gRB1 26 knockdown promoted marker of proliferation Ki-67 (MKi67) expression at 96 h (P < 0.05), 27 and that overexpression of gRB1 reduced MKi67 expression at 72 h (P < 0.05). Together, our 28 study demonstrated that gRB1 inhibited preadipocyte proliferation at least in part by 29 inhibiting the G1 to S phase transition. 30 31 32The selection for rapid growth in meat-type chickens has been accompanied by 36 increased abdominal fat deposition [1]. Excessive abdominal fat deposition can decrease feed 37 efficiency and carcass quality, leading to consumer rejection of the meat [2][3][4]. The excessive 38 deposition of abdominal fat is mainly due to the excessive proliferation and differentiation of 39 adipocytes in adipose tissues. Clarifying the genetic mechanisms of proliferation and 40 differentiation of adipocytes will help control the excessive accumulation of abdominal fat. 41In our previous study, we mapped a quantitative trait locus (QTL) with major effects on 42 abdominal fat trait into a 3.7-Mbp (172.6-176.3 Mbp) region in chicken chromosome 1 using 43 an F 2 population of a broiler × layer cross [5]. In this 3.7-Mbp region, five genes, including 44 Retinoblastoma 1 (RB1), were detected [6]. In the examination of genome-wide selection 45 signatures of abdominal fat content with the chicken 60 k SNP chip and the extended 46 haplotype homozygosity (EHH) assay, a number of genes in the significant core regions were 47 detected, and the RB1 gene was detected once again [7]. In addition, polymorphism analysis 48 of the RB1 gene of Northeast Agricultural University broiler lines divergently selected for 49 abdominal fat content (NEAUHLF) indicated that the polymorphisms of RB1 were associated 50 with abdominal fat content (unpublished data). These results suggest that RB1 plays an 51 important role in the deposition of abdominal fat in broiler chickens. However, the mode of 52 action of RB1 in chicken abdominal fat deposition remains unknown, hence, the aim of this 53 study was to analyze the function of RB1 in the proliferation of chicken preadipocytes.54 Materials and methods 55 Ethics statement 56 All animal work was conducted according to the guidelines for the Care and Use of 57 Experimental Animals established by the Ministry of Science and Technology of the People's 4 58 Rep...