Despite the functional complexity of J-proteins, for perspective, it is important to note that the presence of only a J-domain may be sufficient for some cellular functions (). Such is particularly the case, if the domain is localized to a particular site within a cellular compartment (). Such positioning serves to maintain a high local J-domain concentration, thus targeting Hsp70 to particular client proteins at these sites, without need for direct J-protein interaction with the client itself ().
J-protein function with or without client binding
In the simplest cases, a J-protein consists of little besides the J-domain and sequences required for localization. For example, via a single transmembrane domain the J-domain of scHlj1 is positioned at the cytosolic face of the ER membrane, where it recruits soluble cytosolic Hsp70s to assist in the degradation of proteins exiting the ER (that is ER-associated degradation, ERAD)34.
J-proteins tethered near the polypeptide exit site of the ribosome35
are another example of Hsp70 recruitment to a site having a high concentration of clients (). In this case positioning ensuring prevention of nascent chain aggregation and facilitating folding. Fungi have a specialized ribosome-associated Hsp70, independently tethered to the ribosome36
; however, higher eukaryotes rely on the ribosome associated J-protein (DNAJC2) for ribosomal recruitment of the soluble hHSPA8 (also known as Hsc70), which itself has no intrinsic affinity for the ribosome37, 38
Example of tethering of J-proteins to site of action
A J-domain positioned at the translocon of the inner mitochondrial membrane responsible for translocating polypeptides from the cytosol into the matrix is another example of a “minimal J-protein”. The Hsp70 machinery, of which this J-protein (scPam18 or hDNAJC19) is a part, is not involved in protein folding, rather it drives the movement of polypeptides through the translocon. However, the core Hsp70 machinery, scPam18/hDNAJC19, the mitochondrial Hsp70 (scSsc1/hHSPA9) and the NEF (scMge1/hBAP), that forms this “import motor” follows the basic biochemical rules for Hsp70 machines described above. In the process of driving polypeptide import39
: (1) Hsp70 binds to exposed hydrophobic sequences in unfolded translocating polypeptide; (2) the J-protein stimulating its ATPase activity to enhance interaction with the client and (3) the NEF effecting nucleotide release, and thus dissociation of the client. In addition to the matrix-localized J-domain, scPam18/hDNAJC19 consists of only a transmembrane domain, which serves to localize it in the mitochondrial inner membrane, and a short intermembrane space domain, which directly interacts with the translocon40, 41
. Ssc1/HSPA9 is independently localized to the translocon42, 43
. Thus in this case, and others in which the Hsp70 partner is localized (such as fungal ribosome-associated Hsp70s) the tethered J-domain is not necessary for recruitment of Hsp70 per se. Beyond resulting in high local concentrations of both components, dual localization also serves to precisely position the J-domain juxtaposition to Hsp70 () for optimal ATPase stimulation. This precise positioning likely results in exquisite modulation of the interaction of Hsp70 with client protein (i.e. translocating polypeptide in the case of the import motor).
Remarkably, for some cellular functions, the J-domain - that is the ability to stimulate Hsp70’s ATPase activity - may be sufficient, even without sub-compartment localization (). This functional robustness is illustrated by the ability of solely the J-domain of several cytosolic J-proteins to rescue the severe defects caused by the absence of Class I scYdj1, the most abundant J-protein of the yeast cytosol44
. Surprisingly, this rescue occurs even when a J-domain is expressed at normal scYdj1 levels. This and other observations, also underscore the idea that little if any specificity resides in J-domains themselves45
. Perhaps, facilitation of folding of some newly synthesized proteins by cytosolic Hsp70 needs a J-protein only for stimulation of ATP hydrolysis, but not the direct binding to prevent aggregation or increase the probability of its interaction with Hsp70. However, it should be noted that this J-domain rescue of the effects of the absence of Ydj1 is not complete. As discussed below, client protein binding by J-proteins serves many important functions.