Polarized Distribution of Intracellular Components by Class V Myosins in Saccharomyces cerevisiae

ABSTRACT We identified Ypt11p, a rab-type small GTPase, by its functional and two-hybrid interaction with Myo2p, a class V myosin of the budding yeast Saccharomyces cerevisiae. The tail domain of Myo2p was coimmunoprecipitated with Ypt11p, suggesting that Ypt11p forms a complex with Myo2p at its tail domain in vivo. Mutational analysis of YPT11 suggests that Myo2p is a putative effector of Ypt11p. Deletion of YPT11 induced partial delay of mitochondrial transmission to the bud, and overexpression of YPT11 resulted in mitochondrial accumulation in the bud, indicating that Ypt11p acts positively on mitochondrial distribution toward the bud. We isolated two myo2 mutants, myo2-338 and myo2-573, which showed genetic interactions with YPT11. The myo2-573 mutation, identified by a synthetic lethal interaction with ypt11-null, induced a defect in mitochondrial distribution toward the bud, indicating that Myo2p plays a crucial role in polarized distribution of mitochondria. The myo2-338 mutation was identified as the mutation that abolished the effect of overexpressed YPT11, such as the Ypt11p-dependent accumulation of mitochondria in the bud, and the affinity of Myo2p for Ypt11p was reduced. These results indicate that complex formation of Ypt11p with Myo2p accelerates the function of Myo2p for mitochondrial distribution toward the bud.

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Faculty Opinions recommendation of The class V myosin motor protein, Myo2, plays a major role in mitochondrial motility in Saccharomyces cerevisiae.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1104900.562002 ◽

2008 ◽

Author(s):

Lois Weisman

Keyword(s):

Saccharomyces Cerevisiae ◽

Motor Protein ◽

Class V ◽

Myosin Motor ◽

Mitochondrial Motility

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The class V myosin motor protein, Myo2, plays a major role in mitochondrial motility in Saccharomyces cerevisiae

The Journal of Cell Biology ◽

10.1083/jcb.200709099 ◽

2008 ◽

Vol 181 (1) ◽

pp. 119-130 ◽

Cited By ~ 72

Author(s):

Katrin Altmann ◽

Martina Frank ◽

Daniel Neumann ◽

Stefan Jakobs ◽

Benedikt Westermann

Keyword(s):

Saccharomyces Cerevisiae ◽

Actin Cytoskeleton ◽

Molecular Mechanisms ◽

Budding Yeast ◽

Motor Protein ◽

Mitochondrial Morphology ◽

Direct Role ◽

Class V ◽

Myosin Motor ◽

Mitochondrial Motility

The actin cytoskeleton is essential for polarized, bud-directed movement of cellular membranes in Saccharomyces cerevisiae and thus ensures accurate inheritance of organelles during cell division. Also, mitochondrial distribution and inheritance depend on the actin cytoskeleton, though the precise molecular mechanisms are unknown. Here, we establish the class V myosin motor protein, Myo2, as an important mediator of mitochondrial motility in budding yeast. We found that mutants with abnormal expression levels of Myo2 or its associated light chain, Mlc1, exhibit aberrant mitochondrial morphology and loss of mitochondrial DNA. Specific mutations in the globular tail of Myo2 lead to aggregation of mitochondria in the mother cell. Isolated mitochondria lacking functional Myo2 are severely impaired in their capacity to bind to actin filaments in vitro. Time-resolved fluorescence microscopy revealed a block of bud-directed anterograde mitochondrial movement in cargo binding–defective myo2 mutant cells. We conclude that Myo2 plays an important and direct role for mitochondrial motility and inheritance in budding yeast.

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Pex3 peroxisome biogenesis proteins function in peroxisome inheritance as class V myosin receptors

The Journal of Cell Biology ◽

10.1083/jcb.200902117 ◽

2009 ◽

Vol 187 (2) ◽

pp. 233-246 ◽

Cited By ~ 35

Author(s):

Jinlan Chang ◽

Fred D. Mast ◽

Andrei Fagarasanu ◽

Dorian A. Rachubinski ◽

Gary A. Eitzen ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Mother Cell ◽

Direct Interaction ◽

Integral Membrane Proteins ◽

Eukaryotic Cells ◽

General Mechanism ◽

Peroxisome Biogenesis ◽

Class V ◽

Established Role ◽

In Cells

In Saccharomyces cerevisiae, peroxisomal inheritance from mother cell to bud is conducted by the class V myosin motor, Myo2p. However, homologues of S. cerevisiae Myo2p peroxisomal receptor, Inp2p, are not readily identifiable outside the Saccharomycetaceae family. Here, we demonstrate an unexpected role for Pex3 proteins in peroxisome inheritance. Both Pex3p and Pex3Bp are peroxisomal integral membrane proteins that function as peroxisomal receptors for class V myosin through direct interaction with the myosin globular tail. In cells lacking Pex3Bp, peroxisomes are preferentially retained by the mother cell, whereas most peroxisomes gather and are transferred en masse to the bud in cells overexpressing Pex3Bp or Pex3p. Our results reveal an unprecedented role for members of the Pex3 protein family in peroxisome motility and inheritance in addition to their well-established role in peroxisome biogenesis at the endoplasmic reticulum. Our results point to a temporal link between peroxisome formation and inheritance and delineate a general mechanism of peroxisome inheritance in eukaryotic cells.

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Identification of MYO4, a second class V myosin gene in yeast

Journal of Cell Science ◽

10.1242/jcs.107.4.1055 ◽

1994 ◽

Vol 107 (4) ◽

pp. 1055-1064 ◽

Cited By ~ 12

Author(s):

B.K. Haarer ◽

A. Petzold ◽

S.H. Lillie ◽

S.S. Brown

Keyword(s):

Saccharomyces Cerevisiae ◽

Binding Sites ◽

Coiled Coil ◽

Neck Region ◽

Actin Gene ◽

Yeast Saccharomyces Cerevisiae ◽

Class V ◽

Calmodulin Binding ◽

Coiled Coil Domain ◽

Overexpression Phenotype

We have isolated a fourth myosin gene (MYO4) in yeast (Saccharomyces cerevisiae). MYO4 encodes a approximately 170 kDa (1471 amino acid) class V myosin, using the classification devised by Cheney et al. (1993a; Cell Motil. Cytoskel. 24, 215–223); the motor domain is followed by a neck region containing six putative calmodulin-binding sites and a tail with a short potential ‘coiled-coil’ domain. A comparison with other myosins in GenBank reveals that Myo4 protein is most closely related to the yeast Myo2 protein, another class V myosin. Deletion of MYO4 produces no detectable phenotype, either alone or in conjunction with mutations in myo2 or other myosin genes, the actin gene, or secretory genes. However, overexpression of MYO4 or MYO2 results in several morphological abnormalities, including the formation of short strings of unseparated cells in diploid strains, or clusters of cells in haploid strains. Alterations of MYO4 or MYO2 indicate that neither the motor domains nor tails of these myosins are required to confer the overexpression phenotype, whereas the neck region may be required. Although this phenotype is similar to that seen upon MYO1 deletion, we provide evidence that the overexpression of Myo4p or Myo2p is not simply interfering with Myo1p function.

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Multiple Myo4 motors enhance ASH1 mRNA transport in Saccharomyces cerevisiae

The Journal of Cell Biology ◽

10.1083/jcb.200912011 ◽

2010 ◽

Vol 189 (4) ◽

pp. 755-767 ◽

Cited By ~ 36

Author(s):

Sunglan Chung ◽

Peter A. Takizawa

Keyword(s):

Saccharomyces Cerevisiae ◽

Rna Binding ◽

Purification Method ◽

Class V ◽

Transport Unit ◽

Multiple Copies ◽

Ash1 Mrna ◽

Localization Element

In Saccharomyces cerevisiae, ASH1 mRNA is transported to the bud tip by the class V myosin Myo4. In vivo, Myo4 moves RNA in a rapid and continuous fashion, but in vitro Myo4 is a nonprocessive, monomeric motor that forms a complex with She3. To understand how nonprocessive motors generate continuous transport, we used a novel purification method to show that Myo4, She3, and the RNA-binding protein She2 are the sole major components of an active ribonucleoprotein transport unit. We demonstrate that a single localization element contains multiple copies of Myo4 and a tetramer of She2, which suggests that She2 may recruit multiple motors to an RNA. Furthermore, we show that increasing the number of Myo4–She3 molecules bound to ASH1 RNA in the absence of She2 increases the efficiency of RNA transport to the bud. Our data suggest that multiple, nonprocessive Myo4 motors can generate continuous transport of mRNA to the bud tip.

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Myo4p is a monomeric myosin with motility uniquely adapted to transport mRNA

The Journal of Cell Biology ◽

10.1083/jcb.200707080 ◽

2007 ◽

Vol 178 (7) ◽

pp. 1193-1206 ◽

Cited By ~ 43

Author(s):

Brian D. Dunn ◽

Takeshi Sakamoto ◽

Myoung-Soon S. Hong ◽

James R. Sellers ◽

Peter A. Takizawa

Keyword(s):

Saccharomyces Cerevisiae ◽

Actin Filament ◽

Motor Domain ◽

Secretory Vesicles ◽

Oligomeric State ◽

Yeast Saccharomyces Cerevisiae ◽

Class V ◽

Ash1 Mrna ◽

Motor Density

The yeast Saccharomyces cerevisiae uses two class V myosins to transport cellular material into the bud: Myo2p moves secretory vesicles and organelles, whereas Myo4p transports mRNA. To understand how Myo2p and Myo4p are adapted to transport physically distinct cargos, we characterize Myo2p and Myo4p in yeast extracts, purify active Myo2p and Myo4p from yeast lysates, and analyze their motility. We find several striking differences between Myo2p and Myo4p. First, Myo2p forms a dimer, whereas Myo4p is a monomer. Second, Myo4p generates higher actin filament velocity at lower motor density. Third, single molecules of Myo2p are weakly processive, whereas individual Myo4p motors are nonprocessive. Finally, Myo4p self-assembles into multi-motor complexes capable of processive motility. We show that the unique motility of Myo4p is not due to its motor domain and that the motor domain of Myo2p can transport ASH1 mRNA in vivo. Our results suggest that the oligomeric state of Myo4p is important for its motility and ability to transport mRNA.

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