Myo1 localizes to phagosomes, some of which traffic to the nucleus in a Myo1-dependent manner in Tetrahymena thermophila.
Myo1 is one of 13 myosins in Tetrahymena thermophila. Initially, twelve of the myosins in Tetrahymena were assigned to Class XX in the myosin superfamily but recently re-assigned to a subclass within Class XIV. In a previous study, we reported that genomic knockout of MYO1 affected phagocytosis and macronuclear amitosis. These two phenotypes have appeared disparate because a possible mechanism linking phagocytosis and amitosis was unknown. In the present study, Myo1 localization was investigated in order to further link machinery for phagocytosis and amitosis. Antibodies directed against the Myo1 motor domain detected an immunospecific polypeptide at 175-180 kDa on immunoblots of wild-type proteins. The 175-180 kDa polypeptide was not detected on immunoblots of proteins from the knockout strain. For immunofluorescence microscopy, cells were allowed to internalize fluorescent beads as markers for phagosomes. In wild-type cells, anti-Myo1 and anti-actin antibodies co-localized to the periphery of phagosomes and the macronucleus. In the MYO1-knockout strain only background fluorescence was observed with anti-Myo1 antibody. Confocal x-z series through macronuclei revealed fluorescent beads within the nucleoplasm. Statistical analysis showed a significant difference between the mean distributions of fluorescent beads in the nucleoplasm of wild-type and MYO1-knockout cells. A fluorescent dye was used to label plasma membrane in living cells. Dye-labeled vacuoles trafficked to the macronucleus. Trafficking of phagosomes to the macronucleus in a myosin-dependent manner is a novel finding and a possible mechanism for targeting myosin and actin to the nucleus.
Hosein
Roland E
RE
Department of Biology, Brooklyn College of the City University of New York, Brooklyn 11210, New York, USA.
Gavin
R H
RH
D000199:Actins / Q000032:analysis; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002467:Cell Nucleus / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D018995:Myosin Heavy Chains / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000276:immunology; D010587:Phagocytosis; D010588:Phagosomes / Q000737:chemistry* / Q000502:physiology*; D015800:Protozoan Proteins / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D016808:Tetrahymena thermophila / Q000502:physiology* / Q000648:ultrastructure
D000199:Actins; D000911:Antibodies, Monoclonal; C491829:MYO1 protein, protozoan; D010455:Peptides; D015800:Protozoan Proteins; D018995:Myosin Heavy Chains