Small GTPase Rah/Rab34 is associated with membrane ruffles and macropinosomes and promotes macropinosome formation.
Macropinocytosis is an efficient process for the uptake of nutrients and solute macromolecules into cells from the external environment. Macropinosomes, which are surrounded by actin, are formed from the cell surface membrane ruffles and migrate toward the cell center. We have cloned the entire coding sequence of a member of the Rab family small GTPases, Rah/Rab34. It lacked a consensus sequence for GTP-binding/GTPase domain. Although wild-type Rah exhibited extremely low GTPase activity in vitro, it exerted appreciable GTPase activity in vivo. In fibroblasts, Rah was colocalized with actin to the membrane ruffles and membranes of relatively large vesicles adjacent to the ruffles. These vesicles were identified as macropinosomes on the basis of several criteria. Rah and Rab5 coexisted in some, but not all, macropinosomes. Rah was predominantly associated with nascent macropinosomes, whereas Rab5 was present in endosomes at later stages. The number of macropinosomes in the cells overexpressing Rah increased about 2-fold. The formation of macropinosomes by the treatment of platelet-derived growth factor or phorbol ester was also facilitated by Rah but suppressed by a dominant-negative Rah. Rah-promoted macropinosome formation was retarded by dominant-negative mutants of Rac1 and WAVE2, which are essential for membrane ruffling. These results imply that Rah is required for efficient macropinosome formation from the membrane ruffles.
Sun
Peng
P
Department of Biology, Faculty of Science, and Graduate School of Science and Technology, Chiba University, Yayoicho, Inageku, Japan.
Yamamoto
Hironobu
H
Suetsugu
Shiro
S
Miki
Hiroaki
H
Takenawa
Tadaomi
T
Endo
Takeshi
T
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D011992:Endosomes / Q000378:metabolism; D020558:GTP Phosphohydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007425:Intracellular Membranes / Q000201:enzymology; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D010873:Pinocytosis; D017386:Sequence Homology, Amino Acid
D017931:DNA Primers; D020558:GTP Phosphohydrolases