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Heterotrimeric kinesin II is the microtubule motor protein responsible for pigment dispersion in Xenopus melanophores.

Tuma MC, Zill A, Le Bot N, Vernos I, Gelfand V - J. Cell Biol. (1998)

Bottom Line: Natl.Furthermore, microinjection of melanophores with SUK4, a function-blocking kinesin antibody, inhibited dispersion of lysosomes but had no effect on melanosome transport.We conclude that melanosome dispersion is powered by kinesin II and not by conventional kinesin.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Structural Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.

ABSTRACT
Melanophores move pigment organelles (melanosomes) from the cell center to the periphery and vice-versa. These bidirectional movements require cytoplasmic microtubules and microfilaments and depend on the function of microtubule motors and a myosin. Earlier we found that melanosomes purified from Xenopus melanophores contain the plus end microtubule motor kinesin II, indicating that it may be involved in dispersion (Rogers, S.L., I.S. Tint, P.C. Fanapour, and V.I. Gelfand. 1997. Proc. Natl. Acad. Sci. USA. 94: 3720-3725). Here, we generated a dominant-negative construct encoding green fluorescent protein fused to the stalk-tail region of Xenopus kinesin-like protein 3 (Xklp3), the 95-kD motor subunit of Xenopus kinesin II, and introduced it into melanophores. Overexpression of the fusion protein inhibited pigment dispersion but had no effect on aggregation. To control for the specificity of this effect, we studied the kinesin-dependent movement of lysosomes. Neither dispersion of lysosomes in acidic conditions nor their clustering under alkaline conditions was affected by the mutant Xklp3. Furthermore, microinjection of melanophores with SUK4, a function-blocking kinesin antibody, inhibited dispersion of lysosomes but had no effect on melanosome transport. We conclude that melanosome dispersion is powered by kinesin II and not by conventional kinesin. This paper demonstrates that kinesin II moves membrane-bound organelles.

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Distribution of microtubules is not affected by expression of headless Xklp3. Immunofluorescence staining with an antitubulin primary antibody followed by a Texas red secondary in  a melanophore overexpressing the headless Xklp3. Microtubules  radiate from the cell center, forming a highly organized polar array. The inset is a fluorescence image taken at the green channel,  showing that the cell in the large picture expresses the EGFP-headless Xklp3 fusion protein. Bar, 20 μm.
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Figure 4: Distribution of microtubules is not affected by expression of headless Xklp3. Immunofluorescence staining with an antitubulin primary antibody followed by a Texas red secondary in a melanophore overexpressing the headless Xklp3. Microtubules radiate from the cell center, forming a highly organized polar array. The inset is a fluorescence image taken at the green channel, showing that the cell in the large picture expresses the EGFP-headless Xklp3 fusion protein. Bar, 20 μm.

Mentions: Since pigment aggregation in melanophores depends on microtubules and transfected cells aggregate their pigment normally, microtubules are most likely not affected by overexpression of headless Xklp3. To directly confirm this, microtubules were stained with an antitubulin antibody. Cells overexpressing headless Xklp3 showed normal density and pattern of microtubule organization, with individual microtubules radiating from the cell center and forming a highly organized polar array (Fig. 4). Hence, overexpression of headless Xklp3 specifically interfered with the plus end motility involved in dispersion, but affected neither microtubule organization nor pigment aggregation. Taken together, these results show a direct role of kinesin II in melanosome dispersion.


Heterotrimeric kinesin II is the microtubule motor protein responsible for pigment dispersion in Xenopus melanophores.

Tuma MC, Zill A, Le Bot N, Vernos I, Gelfand V - J. Cell Biol. (1998)

Distribution of microtubules is not affected by expression of headless Xklp3. Immunofluorescence staining with an antitubulin primary antibody followed by a Texas red secondary in  a melanophore overexpressing the headless Xklp3. Microtubules  radiate from the cell center, forming a highly organized polar array. The inset is a fluorescence image taken at the green channel,  showing that the cell in the large picture expresses the EGFP-headless Xklp3 fusion protein. Bar, 20 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2132968&req=5

Figure 4: Distribution of microtubules is not affected by expression of headless Xklp3. Immunofluorescence staining with an antitubulin primary antibody followed by a Texas red secondary in a melanophore overexpressing the headless Xklp3. Microtubules radiate from the cell center, forming a highly organized polar array. The inset is a fluorescence image taken at the green channel, showing that the cell in the large picture expresses the EGFP-headless Xklp3 fusion protein. Bar, 20 μm.
Mentions: Since pigment aggregation in melanophores depends on microtubules and transfected cells aggregate their pigment normally, microtubules are most likely not affected by overexpression of headless Xklp3. To directly confirm this, microtubules were stained with an antitubulin antibody. Cells overexpressing headless Xklp3 showed normal density and pattern of microtubule organization, with individual microtubules radiating from the cell center and forming a highly organized polar array (Fig. 4). Hence, overexpression of headless Xklp3 specifically interfered with the plus end motility involved in dispersion, but affected neither microtubule organization nor pigment aggregation. Taken together, these results show a direct role of kinesin II in melanosome dispersion.

Bottom Line: Natl.Furthermore, microinjection of melanophores with SUK4, a function-blocking kinesin antibody, inhibited dispersion of lysosomes but had no effect on melanosome transport.We conclude that melanosome dispersion is powered by kinesin II and not by conventional kinesin.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Structural Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.

ABSTRACT
Melanophores move pigment organelles (melanosomes) from the cell center to the periphery and vice-versa. These bidirectional movements require cytoplasmic microtubules and microfilaments and depend on the function of microtubule motors and a myosin. Earlier we found that melanosomes purified from Xenopus melanophores contain the plus end microtubule motor kinesin II, indicating that it may be involved in dispersion (Rogers, S.L., I.S. Tint, P.C. Fanapour, and V.I. Gelfand. 1997. Proc. Natl. Acad. Sci. USA. 94: 3720-3725). Here, we generated a dominant-negative construct encoding green fluorescent protein fused to the stalk-tail region of Xenopus kinesin-like protein 3 (Xklp3), the 95-kD motor subunit of Xenopus kinesin II, and introduced it into melanophores. Overexpression of the fusion protein inhibited pigment dispersion but had no effect on aggregation. To control for the specificity of this effect, we studied the kinesin-dependent movement of lysosomes. Neither dispersion of lysosomes in acidic conditions nor their clustering under alkaline conditions was affected by the mutant Xklp3. Furthermore, microinjection of melanophores with SUK4, a function-blocking kinesin antibody, inhibited dispersion of lysosomes but had no effect on melanosome transport. We conclude that melanosome dispersion is powered by kinesin II and not by conventional kinesin. This paper demonstrates that kinesin II moves membrane-bound organelles.

Show MeSH
Related in: MedlinePlus