Identification of multiple RNA features that influence CCR4 deadenylation activity

被引:44
作者
Viswanathan, P [1 ]
Chen, JJ [1 ]
Chiang, YC [1 ]
Denis, CL [1 ]
机构
[1] Univ New Hampshire, Dept Biochem & Mol Biol, Durham, NH 03824 USA
关键词
D O I
10.1074/jbc.M211794200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The CCR4 family proteins are 3'-5'-deadenylases that function in the first step of the degradation of poly(A) mRNA. Here we report the purification to homogeneity of the yeast CCR4 protein and the analysis of its substrate specificities. CCR4 deadenylated a 7N+23A substrate (seven nucleotides followed by 23 A residues) in a distributive manner. Only small differences in CCR4 activity for different A length substrates were observed until only 1 A residue remained. Correspondingly, the K-m for a 25N+2A substrate was found to be at least 20-fold lower than that for a 26N+1A substrate, although their V-max values differed by only 2-fold. In addition, the total length of the RNA was found to contribute to CCR4 activity: up to 17 nucleotides (not necessarily poly(A)) could be recognized by CCR4. Poly(U), poly(C), and poly(G) were also found to be 12-30-fold better inhibitors of CCR4 compared with poly(A), supporting the observation that CCR4 contains a non-poly(A)-specific binding site. Surprisingly, even longer substrates (greater than or equal to45 nucleotides) stimulated CCR4 to become a processive enzyme, suggesting that CCR4 undergoes an additional transition in the presence of such substrates. CCR4 also displayed no difference in its activity with capped or uncapped RNA substrates. These results indicate that CCR4 recognition of its RNA substrates involves several features of the RNA that could be sites in vivo for controlling the rate of specific mRNA deadenylation.
引用
收藏
页码:14949 / 14955
页数:7
相关论文
共 23 条
[1]   INVITRO DEADENYLATION OF MAMMALIAN MESSENGER-RNA BY A HELA-CELL 3' EXONUCLEASE [J].
ASTROM, J ;
ASTROM, A ;
VIRTANEN, A .
EMBO JOURNAL, 1991, 10 (10) :3067-3071
[2]  
ASTROM J, 1992, J BIOL CHEM, V267, P18154
[3]   Computational modeling of eukaryotic mRNA turnover [J].
Cao, D ;
Parker, R .
RNA, 2001, 7 (09) :1192-1212
[4]   CCR4, a 3′-5′ poly(A) RNA and ssDNA exonuclease, is the catalytic component of the cytoplasmic deadenylase [J].
Chen, JJ ;
Chiang, YC ;
Denis, CL .
EMBO JOURNAL, 2002, 21 (06) :1414-1426
[5]   Mechanism and regulation of mRNA polyadenylation [J].
Colgan, DF ;
Manley, JL .
GENES & DEVELOPMENT, 1997, 11 (21) :2755-2766
[6]   The yeast POP2 gene encodes a nuclease involved in mRNA deadenylation [J].
Daugeron, MC ;
Mauxion, F ;
Séraphin, B .
NUCLEIC ACIDS RESEARCH, 2001, 29 (12) :2448-2455
[7]   MECHANISMS OF MESSENGER-RNA DEGRADATION IN EUKARYOTES [J].
DECKER, CJ ;
PARKER, R .
TRENDS IN BIOCHEMICAL SCIENCES, 1994, 19 (08) :336-340
[8]   A TURNOVER PATHWAY FOR BOTH STABLE AND UNSTABLE MESSENGER-RNAS IN YEAST - EVIDENCE FOR A REQUIREMENT FOR DEADENYLATION [J].
DECKER, CJ ;
PARKER, R .
GENES & DEVELOPMENT, 1993, 7 (08) :1632-1643
[9]   Functionally unrelated signalling proteins contain a fold similar to Mg2+-dependent endonucleases [J].
Dlakic, M .
TRENDS IN BIOCHEMICAL SCIENCES, 2000, 25 (06) :272-273
[10]   CCR4 IS A GLUCOSE-REGULATED TRANSCRIPTION FACTOR WHOSE LEUCINE-RICH REPEAT BINDS SEVERAL PROTEINS IMPORTANT FOR PLACING CCR4 IN ITS PROPER PROMOTER CONTEXT [J].
DRAPER, MP ;
LIU, HY ;
NELSBACH, AH ;
MOSLEY, SP ;
DENIS, CL .
MOLECULAR AND CELLULAR BIOLOGY, 1994, 14 (07) :4522-4531