Misfolded proteins in the endoplasmic reticulum (ER) are dislocated out of the 
ER to the cytosol, polyubiquitinated, and degraded by the ubiquitin-proteasome 
system in a process collectively termed ER-associated degradation (ERAD). Recent 
studies have established that a mammalian ER-localized transmembrane J-protein, 
DNAJB12, cooperates with Hsc70, a cytosolic Hsp70 family member, to promote the 
ERAD of misfolded membrane proteins. Interestingly, mammalian genomes have 
another J-protein called DNAJB14 that shows a high sequence similarity to 
DNAJB12. Yet, very little was known about this protein. Here, we report the 
characterization of DNAJB14. Immunofluorescence study and protease protection 
assay showed that, like DNAJB12, DNAJB14 is an ER-localized, single 
membrane-spanning J-protein with its J-domain facing the cytosol. We used 
co-immunoprecipitation assay to find that DNAJB14 can also specifically bind 
Hsc70 via its J-domain to recruit this chaperone to ER membrane. Remarkably, the 
overexpression of DNAJB14 accelerated the degradation of misfolded membrane 
proteins including a mutant of cystic fibrosis transmembrane conductance 
regulator (CFTRΔF508), but not that of a misfolded luminal protein. Furthermore, 
the DNAJB14-dependent degradation of CFTRΔF508 was compromised by MG132, a 
proteasome inhibitor, indicating that DNAJB14 can enhance the degradation of a 
misfolded membrane protein using the ubiquitin-proteasome system. Thus, the 
mammalian ER possesses two analogous J-proteins (DNAJB14 and DNAJB12) that both 
can promote the ERAD of misfolded transmembrane proteins. Compared with DNAJB12 
mRNA that was widely expressed in mouse tissues, DNAJB14 mRNA was expressed more 
weakly, being most abundant in testis, implying its specific role in this 
tissue.