Energetic components VirD4, VirB11 and VirB4 mediate early DNA transfer reactions required for bacterial type IV secretion

被引:170
作者
Atmakuri, K [1 ]
Cascales, E [1 ]
Christie, PJ [1 ]
机构
[1] Univ Texas, Med Sch Houston, Dept Microbiol & Mol Genet, Houston, TX 77030 USA
关键词
D O I
10.1111/j.1365-2958.2004.04345.x
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
Bacteria use type IV secretion systems (T4SS) to translocate DNA (T-DNA) and protein substrates across the cell envelope. By transfer DNA immunoprecipitation (TrIP), we recently showed that T-DNA translocates through the Agrobacterium tumefaciens VirB/D4 T4SS by forming close contacts sequentially with the VirD4 receptor, VirB11 ATPase, the inner membrane subunits VirB6 and VirB8 and, finally, VirB2 pilin and VirB9. Here, by TrIP, we show that nucleoside triphosphate binding site (Walker A motif) mutations do not disrupt VirD4 substrate binding or transfer to VirB11, suggesting that these early reactions proceed independently of ATP binding or hydrolysis. In contrast, VirD4, VirB11 and VirB4 Walker A mutations each arrest substrate transfer to VirB6 and VirB8, suggesting that these subunits energize this transfer reaction by an ATP-dependent mechanism. By co-immunoprecipitation, we supply evidence for VirD4 interactions with VirB4 and VirB11 independently of other T4SS subunits or intact Walker A motifs, and with the bitopic inner membrane subunit VirB10. We reconstituted substrate transfer from VirD4 to VirB11 and to VirB6 and VirB8 by co-synthesis of previously identified 'core' components of the VirB/D4 T4SS. Our findings define genetic requirements for DNA substrate binding and the early transfer reactions of a bacterial type IV translocation pathway.
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收藏
页码:1199 / 1211
页数:13
相关论文
共 51 条
[1]   VirE2, a type IV secretion substrate, interacts with the VirD4 transfer protein at cell poles of Agrobacterium tumefaciens [J].
Atmakuri, K ;
Ding, ZY ;
Christie, PJ .
MOLECULAR MICROBIOLOGY, 2003, 49 (06) :1699-1713
[2]   GENETIC COMPLEMENTATION ANALYSIS OF THE AGROBACTERIUM-TUMEFACIENS VIRB OPERON - VIRB2 THROUGH VIRB11 ARE ESSENTIAL VIRULENCE GENES [J].
BERGER, BR ;
CHRISTIE, PJ .
JOURNAL OF BACTERIOLOGY, 1994, 176 (12) :3646-3660
[3]   THE AGROBACTERIUM-TUMEFACIENS VIRB4 GENE-PRODUCT IS AN ESSENTIAL VIRULENCE PROTEIN REQUIRING AN INTACT NUCLEOSIDE TRIPHOSPHATE-BINDING DOMAIN [J].
BERGER, BR ;
CHRISTIE, PJ .
JOURNAL OF BACTERIOLOGY, 1993, 175 (06) :1723-1734
[4]   INHIBITION OF VIRB-MEDIATED TRANSFER OF DIVERSE SUBSTRATES FROM AGROBACTERIUM-TUMEFACIENS BY THE INCQ PLASMID RSF1010 [J].
BINNS, AN ;
BEAUPRE, CE ;
DALE, EM .
JOURNAL OF BACTERIOLOGY, 1995, 177 (17) :4890-4899
[5]   Genetic evidence of a coupling role for the TraG protein family in bacterial conjugation [J].
Cabezon, E ;
Sastre, JI ;
delaCruz, F .
MOLECULAR & GENERAL GENETICS, 1997, 254 (04) :400-406
[6]   REQUIREMENTS FOR MOBILIZATION OF PLASMIDS RSF1010 AND COLE1 BY THE INCW PLASMID-R388 - TRWB AND RP4-TRAG ARE INTERCHANGEABLE [J].
CABEZON, E ;
LANKA, E ;
DELACRUZ, F .
JOURNAL OF BACTERIOLOGY, 1994, 176 (14) :4455-4458
[7]   Definition of a bacterial type IV secretion pathway for a DNA substrate [J].
Cascales, E ;
Christie, PJ .
SCIENCE, 2004, 304 (5674) :1170-1173
[8]   The versatile bacterial type IV secretion systems [J].
Cascales, E ;
Christie, PJ .
NATURE REVIEWS MICROBIOLOGY, 2003, 1 (02) :137-149
[9]   CONTROLLED EXPRESSION OF THE TRANSCRIPTIONAL ACTIVATOR GENE VIRG IN AGROBACTERIUM-TUMEFACIENS BY USING THE ESCHERICHIA-COLI LAC PROMOTER [J].
CHEN, CY ;
WINANS, SC .
JOURNAL OF BACTERIOLOGY, 1991, 173 (03) :1139-1144
[10]  
CHRISTIE PJ, 2004, BIOCH BIOPHYS ACTA