Even before the identification of Lmf1, biochemical studies in cld
cells and tissues provided important clues about the role of this protein in lipase expression. Despite the virtual absence of LPL activity in plasma and tissues, lipase mRNA and protein expression appeared normal in cld
mice indicating a defect in the posttranslational attainment of lipase activity (15
). It was also demonstrated that inactive LPL formed high-molecular weight aggregates and was retained within the endoplasmic reticulum (ER) of cld
cells consistent with misfolding and ER quality control preventing the release of inactive lipase (16
). In contrast, serum levels of adipsin were unaffected by cld
, arguing against a global defect in protein folding or secretion (15
). Collectively, these studies suggested that the gene affected by the cld
mutation (i.e. Lmf1
) was a lipase-specific chaperone required for the post-translational maturation of lipases into active enzymes.
Lmf1 is a multi-pass transmembrane protein of the ER with three soluble domains protruding into the ER lumen (). This subcellular localization is consistent with the post-translational effect of Lmf1 on lipases, as these enzymes attain catalytic activity within the ER (17
). Co-immunoprecipitation studies revealed that Lmf1 physically interacts with LPL, HL and EL, but not with pancreatic lipase (PL), a related enzyme whose activity is unaffected by the cld
). Although a lipase interaction site has been mapped to one of the ER-facing loops of Lmf1 (), it is clear that other domains also play a critical role in lipase maturation (19
). For example, cld
and the human Y439X mutation (see below), both of which cause C-terminal truncations of Lmf1, completely abolish the maturation of LPL, even though lipase binding remains unaffected. These results suggest complex interactions between Lmf1 and lipases involving multiple protein domains. The location of Lmf1-interaction site(s) within lipases remains to be identified.
What are the molecular mechanisms underlying the lipase-chaperone function of Lmf1? An important clue in this regard is that Lmf1 deficiency only affects lipases that are active as homodimers (LPL, HL and EL), whereas a monomeric member of the lipase superfamily (PL) is unaffected (20
). This observation suggests that Lmf1 is involved in the assembly of inactive lipase monomers into active dimers and/or the stabilization and maintenance of the homodimer structure. Indeed, the LPL dimer is prone to rapid dissociation and inactivation unless stabilized by factors such as heparin in vitro
, or anchoring proteins (heparan sulfate proteoglycans and Gpihbp1) at the endothelial surface in vivo
). In contrast, active LPL is highly stable within the ER (23
). Thus, it is likely that the dimeric lipase structure is maintained by a stabilizing chaperone in this compartment. We propose that Lmf1 is the ER chaperone that plays this role.