Characterization of orchardgrass p23, a flowering plant Hsp90 cohort protein

Abstract

p23 is a heat shock protein 90 (Hsp90) cochaperone and stabilizes the Hsp90 heterocomplex in mammals and yeast. In this study, we isolated a complementary DNA (cDNA) encoding p23 from orchardgrass (Dgp23) and characterized its functional roles under conditions of thermal stress. Dgp23 is a 911 bp cDNA with an open reading frame predicted to encode a 180 amino acid protein. Northern analysis showed that expression of Dgp23 transcripts was heat inducible. Dgp23 has a well-conserved p23 domain and interacted with an orchardgrass Hsp90 homolog in vivo, like mammalian and yeast p23 homologs. Recombinant Dgp23 is a small acidic protein with a molecular mass of approximately 27 kDa and pI 4.3. Dgp23 was also shown to function as a chaperone protein by suppression of malate dehydrogenase thermal aggregation. Differential scanning calorimetry thermograms indicated that Dgp23 is a heat-stable protein, capable of increasing the T m of lysozyme. Moreover, overexpression of Dgp23 in a yeast p23 homolog deletion strain, Δsba1, increased cell viability. These results suggest that Dgp23 plays a role in thermal stress-tolerance and functions as a co-chaperone of Hsp90 and as a chaperone.

A novel thiol-reductase activity of Arabidopsis YUC6 confers drought tolerance independently of auxin biosynthesis

“…Purified Arabidopsis mitochondrial MDH (EC 1.1.1.37) 68 and purified recombinant wild-type and mutant YUC6 proteins at the indicated molar ratios were incubated in 40 mM HEPES (pH 7.5) buffer at 45 °C. The absorbance at 340 nm, indicating aggregation of MDH, was monitored for 30 min using a Beckman DU-800 spectrophotometer (Beckman Coulter) attached to a thermostatic cell holder assembly 58 60 68 .…”

Heat Shock Proteins in Association with Heat Tolerance in Grasses

“…The HSP20-like chaperone is highly homologous to an HSP20-like chaperone from clover ( Medicago truncatula ) that contains the p23 domain. As p23 is one of the cochaperones of HSP90 and stabilizes the HSP90 heterocomplex [ 98 , 99 ], enhanced expression of this chaperone gene under heat stress also indicated that upregulation of HSP90-related proteins are important for heat tolerance in perennial grasses, as discussed above in the proteomic study. In another study, using the sequence of the HSP70 gene isolated by SSH in A. scabra and the reported sequence of a sHSP ( HSP16 ) gene in A. stolonifera , the expression levels of the two genes were compared between heat-sensitive A. stolonifera and thermal A. scabra (Huang, unpublished data).…”

Expression of five AtHsp90 genes in Saccharomyces cerevisiae reveals functional differences of AtHsp90s under abiotic stresses

“…Deletion studies suggest that human and yeast p23 interacts with both the N-terminal and middle domains of Hsp90␤ in vitro (McLaughlin et al, 2006), whereas plant p23 is not like human and yeast p23, interacting with cytosolic Hsp90 (Krishna and Zhang, 2003). However, in a recent study, it was shown that p23 from orchardgrass could interact with DgHsp90, which is a Hsp90 homolog in orchardgrass located in the ER (Cha et al, 2009). In this study, organellar and cytosolic AtHsp90s showed different interactions with the tested cofactors, cpHsp70, Hsp70, Hsp70t-2, Cyp40, p23 and a substrate protein of NOS, organellar AtHsp90s could interact with most of the tested cofactors, while cytosolic AtHsp90s could not interact with these cofactors (Fig.…”