Actinin-4 is preferentially involved in circular ruffling and macropinocytosis in mouse macrophages: analysis by fluorescence ratio imaging.
We have applied fluorescence ratio imaging to the analysis of an actin-binding protein concentration relative to F-actin in macrophages, in order to explore the role of a novel (alpha)-actinin isoform, actinin-4, relative to that of the classical isoform, actinin-1. Conventional immunofluorescence images showed that both isoforms were enriched in F-actin-rich regions such as cell surface ruffles. However, ratio images further demonstrated that actinin-4 concentrations relative to F-actin were higher in peripheral inward curved ruffles and dorsal circular ruffles, presumed precursor forms of macropinosomes, than in straight linear ruffles, while actinin-1 concentrations were uniform among the different types of ruffles. Macropinosome pulse-labeling and chase experiments indicated that actinin-4 was also closely associated with newly formed macropinosomes and gradually dissociated with their maturation. Consistent with ratio imaging data, macrophages scrape-loaded with anti-actinin-4 showed a more reduced rate of macropinocytosis than those loaded with anti-actinin-1. Altogether, these results indicate that actinin-4 and actinin-1 contribute differently to F-actin dynamics, that actinin-4 is more preferentially involved in early stages of macropinocytosis than actinin-1. A similar redistribution of actinin-4 was also observed during phagocytosis, suggesting that actinin-4 may play the same role in the two mechanistically analogous types of endocytosis, i.e. macropinocytosis and phagocytosis.
Araki
N
N
Department of Anatomy, Kagawa Medical University, Miki, Kagawa 761-0793, Japan. naraki@kms.ac.jp
Hatae
T
T
Yamada
T
T
Hirohashi
S
S
D000185:Actinin / Q000276:immunology / Q000378:metabolism / Q000502:physiology*; D000818:Animals; D000918:Antibody Specificity; D002468:Cell Physiological Phenomena; D002478:Cells, Cultured; D018922:HL-60 Cells; D006801:Humans; D008264:Macrophages / Q000502:physiology* / Q000648:ultrastructure; D051379:Mice; D008809:Mice, Inbred C3H; D008840:Microfilament Proteins; D018613:Microscopy, Confocal / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D010587:Phagocytosis / Q000502:physiology; D010873:Pinocytosis / Q000502:physiology*; D020033:Protein Isoforms / Q000276:immunology / Q000378:metabolism / Q000502:physiology; D020298:U937 Cells
C111453:ACTN4 protein, human; C491752:Actn4 protein, mouse; D008840:Microfilament Proteins; D020033:Protein Isoforms; D000185:Actinin