Actin‐dependent motility of melanosomes from fish retinal pigment epithelial (RPE) cells investigated using in vitro motility assays

McNeil, Eileen; Tacelosky, D.; Basciano, Paul; Biallas, B.; Williams, R. D.; Damiani, P.; Deacon, Sean; Fox, C F; Stewart, Bonnie J.; Petruzzi, N.; Osborn, Charlene P.; Klinger, Katherine W.; Sellers, James R.; Smith, Christina King

doi:10.1002/cm.10179

Cited by 18 publications

(16 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Data in this report are consistent with motor-driven motility of phagosomes, although there is no direct evidence. Velocity of phagosomes in control cells and in both treatment groups is consistent with the velocity of myosin-based motility of melanosomes in fish retinal-pigmented epithelium (RPE) [McNeil et al, 2004], uncoated endocytic vesicles in mammalian RPE [Aschenbrenner et al, 2004], and melanosomes in mammalian RPE [Gibbs et al, 2004]. The asymmetric arrangement of phagosome-associated actin invites comparison with actin-rich rocket tails, but the velocity of phagosome motility is considerably slower than actin-driven motility of phagosomes in mouse macrophages [Zhang et al, 2002], pinosomes in cultured mast cells [Merrifield et al, 1999], and endosomes and lysosomes in Xenopus eggs [Taunton et al, 2000].…”

Section: Discussionsupporting

confidence: 67%

Directed motility of phagosomes inTetrahymena thermophila requires actin and Myo1p, a novel unconventional myosin

Hosein

Williams

Gavin

2005

Cell Motil. Cytoskeleton

View full text Add to dashboard Cite

The phagosome cycle was investigated in Tetrahymena thermophila, which had internalized fluorescent latex beads. Confocal microscopy of cells from a GFP-actin strain revealed actin filaments that extended 3-5 mum from the periphery of fluorescent phagosomes. In GFP-actin cells and in wild-type cells, motility of fluorescent phagosomes was directed from the oral cavity to the posterior end of the cell. Although 60% of fluorescent phagosomes in the MYO1-knockout strain were motile, movement of phagosomes was not directed toward the posterior end of the cell and was random. Forty percent of fluorescent phagosomes in knockout cells were non-motile in contrast to only 20% non-motile phagosomes in wild-type cells. The increased incidence of non-motile phagosomes in the knockout strain could reflect absence of Myo1p as a motor. Another myosin or other molecular motors could power random movement of phagosomes in the MYO1-knockout strain. In latrunculin-treated GFP-actin cells, movement of fluorescent phagosomes was random. Average velocity of random movement of fluorescent phagosomes in the knockout strain and in latrunculin-treated cells was statistically the same as the average velocity (2.0 +/- 1.9 microm/min) of phagosomes in GFP-actin cells. These findings are an indication that dynamic actin and Myo1p are required for directed motility of phagosomes.

show abstract

Section: Discussionsupporting

confidence: 67%

Directed motility of phagosomes inTetrahymena thermophila requires actin and Myo1p, a novel unconventional myosin

Hosein

Williams

Gavin

2005

Cell Motil. Cytoskeleton

View full text Add to dashboard Cite

show abstract

“…Therefore, one possibility myosin II in fish RPE associates directly with melanosomes and translocates them along actin filaments. Although myosin VIIa is enriched on purified melanosomes from fish RPE [McNeil et al, 2004], different rate classes observed in in vitro motility assays in that study suggested the possibility that more than one type of myosin is present on melanosome membranes. Possibly myosin II is one of these other types of myosins on RPE melanosomes.…”

Section: Role Of Myosin II In Melanosome Aggregationmentioning

confidence: 80%

Myosin II and Rho kinase activity are required for melanosome aggregation in fish retinal pigment epithelial cells

Barsoum¹,

King-Smith²

2007

Cell Motil. Cytoskeleton

View full text Add to dashboard Cite

In the retinal pigment epithelium (RPE) of fish, melanosomes (pigment granules) migrate long distances through the cell body into apical projections in the light, and aggregate back into the cell body in the dark. RPE cells can be isolated from the eye, dissociated, and cultured as single cells in vitro. Treatment of isolated RPE cells with cAMP or the phosphatase inhibitor, okadaic acid (OA), stimulates melanosome aggregation, while cAMP or OA washout in the presence of dopamine triggers dispersion. Previous studies have shown that actin filaments are both necessary and sufficient for aggregation and dispersion of melanosomes within apical projections of isolated RPE. The role of myosin II in melanosome motility was investigated using the myosin II inhibitor, blebbistatin, and a specific rho kinase (ROCK) inhibitor, H-1152. Blebbistatin and H-1152 partially blocked melanosome aggregation triggered by cAMP in dissociated, isolated RPE cells and isolated sheets of RPE. In contrast, neither drug affected melanosome dispersion. In cells exposed to either blebbistatin or H-1152, then triggered to aggregate using OA, melanosome aggregation was completely inhibited. These results demonstrate that (1) melanosome aggregation and dispersion occur through different, actin-dependent mechanisms; (2) myosin II and ROCK activity are required for full melanosome aggregation, but not dispersion; (3) partial aggregation that occurred despite myosin II or ROCK inhibition suggests a second component of aggregation that is dependent on cAMP signaling, but independent of ROCK and myosin II.

show abstract

“…We found that myosin Va is not required for the correct localization of RPE melanosomes, however, it was enriched in the subcellular fraction containing melanosomes, suggesting a role in melanosome transport, even though it is not essential for the localization of the melanosomes. The movements of melanosomes in teleost RPE in response to lighting changes requires an intact actin cytoskeleton (King-Smith et al, 1997), and the melanosomes have been shown recently, in elegant motility studies, to contain a plus-end directed myosin(s) (McNeil et al, 2004). Interestingly, however, light-dependent movements of melanosomes still occur in the RPE of mariner zebrafish (Perkins et al, 2004), which lack myosin VIIa (Ernest et al, 2000), indicating that myosin VIIa is not required for these movements.…”

Section: Discussionmentioning

confidence: 99%

Role of myosin VIIa and Rab27a in the motility and localization of RPE melanosomes

Gibbs

Azarian

Lillo

et al. 2004

Journal of Cell Science

135

159

View full text Add to dashboard Cite

Myosin VIIa functions in the outer retina, and loss of this function causes human blindness in Usher syndrome type 1B (USH1B). In mice with mutant Myo7a, melanosomes in the retinal pigmented epithelium (RPE) are distributed abnormally. In this investigation we detected many proteins in RPE cells that could potentially participate in melanosome transport, but of those tested, only myosin VIIa and Rab27a were found to be required for normal distribution. Two other expressed proteins, melanophilin and myosin Va, both of which are required for normal melanosome distribution in melanocytes, were not required in RPE, despite the association of myosin Va with the RPE melanosome fraction. Both myosin VIIa and myosin Va were immunodetected broadly in sections of the RPE, overlapping with a region of apical filamentous actin. Some 70-80% of the myosin VIIa in RPE cells was detected on melanosome membranes by both subcellular fractionation of RPE cells and quantitative immunoelectron microscopy, consistent with a role for myosin VIIa in melanosome motility. Time-lapse microscopy of melanosomes in primary cultures of mouse RPE cells demonstrated that the melanosomes move in a saltatory manner, interrupting slow movements with short bursts of rapid movement (>1 μm/second). In RPE cells from Myo7a-null mice, both the slow and rapid movements still occurred, except that more melanosomes underwent rapid movements, and each movement extended approximately five times longer (and further). Hence, our studies demonstrate the presence of many potential effectors of melanosome motility and localization in the RPE, with a specific requirement for Rab27a and myosin VIIa, which function by transporting and constraining melanosomes within a region of filamentous actin. The presence of two distinct melanosome velocities in both control and Myo7a-null RPE cells suggests the involvement of at least two motors other than myosin VIIa in melanosome motility, most probably, a microtubule motor and myosin Va.

show abstract

Actin‐dependent motility of melanosomes from fish retinal pigment epithelial (RPE) cells investigated using in vitro motility assays

Cited by 18 publications

References 34 publications

Directed motility of phagosomes inTetrahymena thermophila requires actin and Myo1p, a novel unconventional myosin

Directed motility of phagosomes inTetrahymena thermophila requires actin and Myo1p, a novel unconventional myosin

Myosin II and Rho kinase activity are required for melanosome aggregation in fish retinal pigment epithelial cells

Role of myosin VIIa and Rab27a in the motility and localization of RPE melanosomes

Contact Info

Product

Resources

About