Functional characterization of the MENTAL domain.
Human metastatic lymph node (MLN) 64 is composed of two conserved regions. The amino terminus contains a conserved membrane-spanning MENTAL (MLN64 NH(2)-terminal) domain shared with an unique protein called MENTHO (MLN64 NH(2)-terminal domain homologue) and targets the protein to late endosome. The carboxyl-terminal domain is composed of a cholesterol binding steroidogenic acute regulatory-related lipid transfer domain exposed to the cytoplasm. MENTHO overexpression leads to the accumulation of enlarged endosomes. In this study, we show that MLN64 overexpression also induces the formation of enlarged endosomes, an effect that is probably mediated by the MENTAL domain. Using an in vivo photocholesterol binding assay, we find that the MENTAL domain of MLN64 is a cholesterol binding domain. Moreover, glutathione S-transferase pull-down or co-immunoprecipitation experiments demonstrate that this domain mediates homo- and hetero-interaction of MLN64 and MENTHO. In living cells, the expression of paired yellow fluorescent and cyan fluorescent fusion proteins show MENTHO homo-interaction and its interaction with MLN64. These data indicate that within late-endosomal membranes, MLN64 and MENTHO define discrete cholesterol-containing subdomains. The MENTAL domain might serve to maintain cholesterol at the membrane of late endosomes prior to its shuttle to cytoplasmic acceptor(s).
Alpy
Fabien
F
Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Département de Pathologie Moléculaire, UPR 6520 CNRS/U596 INSERM/Université Louis Pasteur, BP10142, 67404 Illkirch, C. U. de Strasbourg, France.
Latchumanan
Vinoth K
VK
Kedinger
Valérie
V
Janoshazi
Agnes
A
Thiele
Christoph
C
Wendling
Corinne
C
Rio
Marie-Christine
MC
Tomasetto
Catherine
C
D002352:Carrier Proteins / Q000737:chemistry / Q000502:physiology*; D011992:Endosomes / Q000737:chemistry / Q000378:metabolism / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000502:physiology*; D010446:Peptide Fragments / Q000737:chemistry / Q000502:physiology*; D017434:Protein Structure, Tertiary
D002352:Carrier Proteins; D008565:Membrane Proteins; D010446:Peptide Fragments; C105684:STARD3 protein, human; C471300:STARD3NL protein, human