Tuesday, June 15, 2010

UMA and MABP domains in receptor endocytosis and endosomal sorting

Elucidating interactions between ESCRT complexes and membrane proteins or lipids is critical to understand endosomal trafficking. We identified two domains with potential significance for this process. The MABP (MVB12-associated domain beta prism domain)found in animal ESCRT-I subunit MVB12, Crag, a regulator of protein sorting, and bacterial pore-forming proteins might mediate novel membrane interactions in trafficking. The UMA domain found in MVB12 and UBAP1/2 helps define a novel class of adaptor domains that might recruit diverse targets to ESCRT-I.

A key aspect of eukaryotic intracellular trafficking is the sorting of cell-surface proteins into multi-vesicular endosomes or bodies (MVBs), which eventually fuse with the lysosome, where they are degraded by lipases and peptidases. This is the primary mechanism for down-regulation of signaling via transmembrane receptors and removal of misfolded or defective membrane proteins. This mechanism is also utilized by several viruses (e.g. HIV-1) to facilitate budding of their virions from the cell-membrane. Studies on animal and fungal models have shown that the process depends on an intricate series of interactions that is initiated via ubiquitination (typically one or more mono-ubiquitinations) of the cytoplasmic tails of membrane proteins by specific E3 ubiquitin (Ub) ligases. The ubiquitinated membrane proteins are then captured into endosomes by the ESCRT system and prevented from being recycled back to the plasma membrane via the retrograde trafficking system. The ESCRT system also folds the endosomal membranes into invaginations that are concentrated in these ubiqutinated targets and catalyzes their abscission into intra-luminal-vesicles inside the endosome. This largely seals the fate of these membrane proteins as targets for lysosomal degradation. The ESCRT system itself comprises of 4 major protein complexes, ESCRT0 to ESCRT-III, which are successively involved in the above-described steps. ESCRT-0 with proteins bearing multiple Ub-binding modules is the primary sensor for the subset of membrane proteins that are ubiquitinated. Both ESCRT-I and ESCRT-II have proteins with a single Ub-binding domain and appear to be the successive recipients of the ubiqutinated cargo initially sensed by ESCRT-0. The ESCRT-II complex also contains lipid-binding modules and is likely to initiate the invagination of the endosomal membrane. ESCRT-III, which includes the conserved AAA+ ATPase VPS4 as a component, mediates the final abscission of the invaginated membrane to form the ILV. In this relay the ESCRT-I complex is a central player, for it acts as a bridge between the initial sensor of the ubiquitinated targets and membrane-binding ESCRT-II.

Domain architectures of the UMA and MABP domains/ Structure of the MABP domain

ESCRT-I is comprised of three major subunits that are conserved between yeast and animals, namely the inactive E2-ligase protein TSG101/VPS23, Vps28 and Vps37. Additionally, both fungal and animal ESCRT-Is contain a fourth subunit termed MVB12; however, the MVB12 subunits from the two lineages were found to be not homologous. The animal MVB12 was shown to be critical for receptor endocy-tosis and also virus release. Given its key role in receptor down-regulation we were interested in understanding if the lack of the homology with the fungal MVB12 might reflect the emergence of novel interactions related to the expansion of receptor-mediated signaling in animals.

Identification of the MABP and UMA domains throws light on two key aspects of vesicular trafficking. Firstly, the MABP domain could be a common denominator in the recognition of specific membrane-associated features by a functionally diverse set of traf-ficking proteins in eukaryotes and bacterial proteins involved in pore formation and cell-wall interaction. The prediction that the diverse metazoan UMA domain proteins are alternative MVB12-like proteins implies that the recruitment of ESCRT-I to endosomal structures could occur via diverse mechanisms, including the poss-ible direct recognition of membranes by the MABP domain, inte-raction with ubiquitinated peptides or other protein-protein interac-tions. The UMA domain is currently only detectable in metazoans and appears unrelated to the yeast MVB12 protein. This is consistent with the vast expansion of diverse signaling receptors such as receptor tyrosine kinases, ion channels and 7TM receptors in the metazoan lineage and indicates the emergence of a dedicated receptor attenuating system that functions via ESCRT-I. Intriguingly, we found that plants (e.g. ArabidopsisAT5G53330) have a conserved protein that has a series of C-terminal UBA domains closely related to those found in UBAP-1/2. While we failed to find statistically significant similarity between the N-terminal region of these plant proteins and the UMA domain, they share a few tantalizing sequence patterns. Hence, it cannot be ruled out that these plant proteins contain a region remotely related to the UMA domain and perform a comparable function in relation to the ESCRT complex.

While certain core components of the ESCRT complex (e.g. VPS4 and MIT domains of ESCRT-III) have been traced to archaea, the MABP domain is not currently found in any archaea. Instead it is found in several bacteria, suggesting that the eukaryotes could have potentially acquired it early in their evolution from a bacterial precursor. Thus, the eukaryotic vesicular trafficking system appears to have been pieced together from different components acquired from both archaeal and bacterial precursors.