熱處理對錦鯉睪丸細胞株之Ubiquitin基因表現之效應

Abstract

Ubiquitin是一個具有76個胺基酸的蛋白質，普遍存在於目前被檢視過的真核生物中；它參與了細胞的多項生理功能，如：蛋白質的分解，染色質結構的維持，細胞週期的控制以及Heat shock反應等。本實驗的目的是在於瞭解錦鯉睪丸細胞株在受熱處理後，Ubiquitin基因表現的情形。以鯉魚單一的Ubiquitin及銜接於其後而含有52個胺基酸之延伸蛋白之串連cDNA（合稱Ubf52）為探針，進行Northern blot解析，結果顯示：有兩種和Ubiquitin有關的基因表現，其mRNA長度分別為0.6Kb及2.8Kb, 0.6Kb的mRNA在正常細胞及熱處理胞中約有穩定表現，而2.8Kb的mRNA則在熱處理細胞中有大量表現。 此外，以帶Ubiquitin基因的探針，篩選用熱處理兩小時的錦鯉睪丸細胞株所建立互補基因庫，找到一個長度為0.6Kb的CDNA，經核酸序列分析及比對，證實為Ubf52 cDNA基因。

Ubiquitin, a 76-amino acid protein, is present in all eukaryotes examined to date. It is involved in many aspects of cellular functions such as: degradation of proteins, maintenance of chromatin structure, progression of cell cycle and heat shock responces, etc. In this report. the expression of ubiquitin gene in colored carp testis cell line (CCT cell line) with heat shock treatment we studied. Cells were treated at 38℃ for various periods and then the total RNA was analyzed by Northern blot. Before heat treatment, only the ubiquitin mRNA of 0.6 Kb was detected. In addition to the 0.6Kb of ubiquitin mRNA found in untreated cells, .a .ubiquitin mRNA of 2,8 Kb was also expressed after heat shock. In order to understand the gene structure of ubiquitin in CCT cell line. probes with ubiquitin DNA were used to screen a cDNA library constructed from CCT cells treated at 38℃ for two-hour. One cDNA clone was isolated. sequenced and shown to encode a ubiquitin fused to carboxyl extension protein of 52 amino acids. The results indicate it is a ubf52 gene.