Identification of a Site of Hsp27 Binding with Hsp27 and αB-Crystallin as Indicated by the Yeast Two-Hybrid System

Liu, Chenghua; Welsh, Michael J.

doi:10.1006/bbrc.1999.0174

Cited by 39 publications

(28 citation statements)

References 32 publications

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“…b The activation of the lacZ reporter gene was determined by ␤-galactosidase activity as described previously (30). The ␤-galactosidase units (U) were calculated as follows: U ϭ (1,000 ϫ OD at 420 nm)/(reaction time [minutes] ϫ OD at 600 nm ϫ concentration factor), where OD is the optical density.…”

Section: Resultsmentioning

confidence: 99%

Type D Retrovirus Gag Polyprotein Interacts with the Cytosolic Chaperonin TRiC

Choi

Park

Chung

et al. 2001

J Virol

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The carboxy terminus-encoding portion of the gag gene of Mason-Pfizer monkey virus (M-PMV), the prototype immunosuppressive primate type D retrovirus, encodes a 36-amino-acid, proline-rich protein domain that, in the mature virion, becomes the p4 capsid protein. The p4 domain has no known role in M-PMV replication. We found that two mutants with premature termination codons that remove half or all of the p4 domain produced lower levels of stable Gag protein and of self-assembled capsids. Interestingly, yeast twohybrid screening revealed that p4 specifically interacted with TCP-1␥, a subunit of the chaperonin TRiC (TCP-1 ring complex). TRiC is a cytosolic chaperonin that is known to be involved in both folding and subunit assembly of a variety of cellular proteins. TCP-1␥ also associated with high specificity with the M-PMV pp24/16-p12 domain and human immunodeficiency virus p6. Moreover, in cells, Gag polyprotein associated with the TRiC chaperonin complex and this association depended on ATP hydrolysis. In the p4 truncation mutants, the Gag-TRiC association was significantly reduced. These results strongly suggest that cytosolic chaperonin TRiC is involved in Gag folding and/or capsid assembly. We propose that TRiC associates transiently with nascent M-PMV Gag molecules to assist in their folding. Consequently, properly folded Gag molecules carry out the intermolecular interactions involved in self-assembly of the immature capsid.

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Section: Resultsmentioning

confidence: 99%

Type D Retrovirus Gag Polyprotein Interacts with the Cytosolic Chaperonin TRiC

Choi

Park

Chung

et al. 2001

J Virol

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“…An interaction between Hsp27 and ␣B-crystallin was demonstrated by the yeast two-hybrid system (27,190). Finally, fluorescence-labeled ␣A-crystallin readily exchanged with ␣B-crystallin and with Hsp27 (31).…”

Section: Hetero-oligomeric Complexesmentioning

confidence: 99%

“…Congruent is the finding that a genetically engineered human ␣B-crystallin comprising only the ␣-crystallin domain forms a dimer (81). Yeast twohybrid system analyses provide genetic evidence for interactions in the ␣-crystallin domain (27,157,168,190,359). The conserved motif A-x-x-x-n-G-v-L (Fig.…”

Section: Regions Responsible For Oligomerizationmentioning

confidence: 99%

α-Crystallin-Type Heat Shock Proteins: Socializing Minichaperones in the Context of a Multichaperone Network

Narberhaus

2002

Microbiol Mol Biol Rev

498

415

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SUMMARY α-Crystallins were originally recognized as proteins contributing to the transparency of the mammalian eye lens. Subsequently, they have been found in many, but not all, members of the Archaea, Bacteria, and Eucarya. Most members of the diverse α-crystallin family have four common structural and functional features: (i) a small monomeric molecular mass between 12 and 43 kDa; (ii) the formation of large oligomeric complexes; (iii) the presence of a moderately conserved central region, the so-called α-crystallin domain; and (iv) molecular chaperone activity. Since α-crystallins are induced by a temperature upshift in many organisms, they are often referred to as small heat shock proteins (sHsps) or, more accurately, α-Hsps. α-Crystallins are integrated into a highly flexible and synergistic multichaperone network evolved to secure protein quality control in the cell. Their chaperone activity is limited to the binding of unfolding intermediates in order to protect them from irreversible aggregation. Productive release and refolding of captured proteins into the native state requires close cooperation with other cellular chaperones. In addition, α-Hsps seem to play an important role in membrane stabilization. The review compiles information on the abundance, sequence conservation, regulation, structure, and function of α-Hsps with an emphasis on the microbial members of this chaperone family.

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“…The basic 600 kDa oligomers are roughly spherical and interact to form larger complexes (Shearstone & Baneyx, 1998). There is no evidence that IbpA and IbpB are assembled into mixed complexes in E. coli cells, although in organisms containing more than one sHsp, different sHsp species form hetero-oligomers, for example : vertebrate αA-αB-crystallin, αB-crystallin-Hsp27 and sHsps from Bradyrhizobium japonicum (HspH\HspB and HspC\HspF) (MacRea, 2000 ;Liu & Welsh, 1999 ;Studer & Narberhaus, 2000). IbpB was also found to form mixed oligomers with HspB and HspH of B. japonicum (Studer & Narberhaus, 2000).…”

Section: Abbreviationsmentioning

confidence: 99%

The Escherichia coli small heat-shock proteins IbpA and IbpB prevent the aggregation of endogenous proteins denatured in vivo during extreme heat shock

et al. 2002

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The roles of the Escherichia coli IbpA and IbpB chaperones in protection of heat-denatured proteins against irreversible aggregation in vivo were investigated. Overproduction of IbpA and IbpB resulted in stabilization of the denatured and reversibly aggregated proteins (the S fraction), which could be isolated from E. coli cells by sucrose gradient centrifugation. This finding is in agreement with the present model of the small heat-shock proteins' function, based mainly on in vitro studies. Deletion of the ibpAB operon resulted in almost twofold increase in protein aggregation and in inactivation of an enzyme (fructose-1,6-biphosphate aldolase) in cells incubated at 50 SC for 4 h, decreased efficiency of the removal of protein aggregates formed during prolonged incubation at 50 SC and affected cell viability at this temperature. IbpA/B proteins were not needed for removal of protein aggregates or for the enzyme protection/renaturation in cells heat shocked at 50 SC for 15 min. These results show that the IbpA/B proteins are required upon an extreme, long-term heat shock. Overproduction of IbpA but not IbpB caused an increase of the level of β-lactamase precursor, which was localized in the S fraction, together with the IbpA protein, which suggests that the unfolded precursor binds to IbpA but not to IbpB. Although in the wild-type cells both E. coli small heatshock proteins are known to localize in the S fraction, only 2 % of total IbpB co-localized with the aggregated proteins in the absence of IbpA, while in the absence of IbpB, the majority of IbpA was present in the aggregates fraction.

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Identification of a Site of Hsp27 Binding with Hsp27 and αB-Crystallin as Indicated by the Yeast Two-Hybrid System

Cited by 39 publications

References 32 publications

Type D Retrovirus Gag Polyprotein Interacts with the Cytosolic Chaperonin TRiC

Type D Retrovirus Gag Polyprotein Interacts with the Cytosolic Chaperonin TRiC

α-Crystallin-Type Heat Shock Proteins: Socializing Minichaperones in the Context of a Multichaperone Network

The Escherichia coli small heat-shock proteins IbpA and IbpB prevent the aggregation of endogenous proteins denatured in vivo during extreme heat shock

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