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Activated mutant NRas(Q61K) drives aberrant melanocyte signaling, survival, and invasiveness via a Rac1-dependent mechanism.

Li A, Ma Y, Jin M, Mason S, Mort RL, Blyth K, Larue L, Sansom OJ, Machesky LM - J. Invest. Dermatol. (2012)

Bottom Line: The Ras oncogene also activates signaling via the related Rho GTPase Rac1, which can mediate growth, survival, and motility signaling.We tested the effects of activated NRas(Q61K) on the proliferation, motility, and invasiveness of melanoblasts and melanocytes in the developing mouse and ex vivo explant culture as well as in a melanoma transplant model.We also show that genetic deletion or pharmacological inhibition of Rac1 in NRas(Q61K) induced melanoma suppresses tumor growth, lymph node spread, and tumor cell invasiveness, suggesting a potential value for Rac1 as a therapeutic target for activated NRas-driven tumor growth and invasiveness.

View Article: PubMed Central - PubMed

Affiliation: The Beatson Institute for Cancer Research, Garscube Estate, Glasgow, UK.

ABSTRACT
Around a fifth of melanomas exhibit an activating mutation in the oncogene NRas that confers constitutive signaling to proliferation and promotes tumor initiation. NRas signals downstream of the major melanocyte tyrosine kinase receptor c-kit and activated NRas results in increased signaling via the extracellular signal-regulated kinase (ERK)/MAPK/ERK kinase/mitogen-activated protein kinase (MAPK) pathways to enhance proliferation. The Ras oncogene also activates signaling via the related Rho GTPase Rac1, which can mediate growth, survival, and motility signaling. We tested the effects of activated NRas(Q61K) on the proliferation, motility, and invasiveness of melanoblasts and melanocytes in the developing mouse and ex vivo explant culture as well as in a melanoma transplant model. We find an important role for Rac1 downstream of NRas(Q61K) in mediating dermal melanocyte survival in vivo in mouse, but surprisingly NRas(Q61K) does not appear to affect melanoblast motility or proliferation during mouse embryogenesis. We also show that genetic deletion or pharmacological inhibition of Rac1 in NRas(Q61K) induced melanoma suppresses tumor growth, lymph node spread, and tumor cell invasiveness, suggesting a potential value for Rac1 as a therapeutic target for activated NRas-driven tumor growth and invasiveness.

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Mice expressing melanocyte lineage NRasQ61K retain pigmentation defects caused by Rac1 deletion. Coat color of dorsal (a) and ventral side (b) of P14 Tyr∷NRasQ61K+/o Rac1 f/f Tyr∷Cre+/o (Rac1 f/f NRas) mouse with Tyr∷NRasQ61K+/o (NRas) control (Ctr) littermate. Typical dorsal white patches Rac1 f/f NRas mouse (arrows). Coat color of forelimb (c) and tail (d) of Rac1 f/f NRas mouse with NRas Ctr littermate. (e) Quantification of white patches on dorsal side of mice. Body weight of mice at P7 (f) and P14 (g); n=6 mice from three different litters. Lower, median, and upper quartile are shown. (h) Ventral skin from P14 NRas Ctr and Rac1 f/f NRas mice with anti-dopachrome tautomerase (melanocytes). Typical cluster of dermal melanocytes in NRas Ctr mouse is shown with an arrow. **P<0.01 compared with Ctr by t-test. Bar=100 μm. NS, not significant.
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fig2: Mice expressing melanocyte lineage NRasQ61K retain pigmentation defects caused by Rac1 deletion. Coat color of dorsal (a) and ventral side (b) of P14 Tyr∷NRasQ61K+/o Rac1 f/f Tyr∷Cre+/o (Rac1 f/f NRas) mouse with Tyr∷NRasQ61K+/o (NRas) control (Ctr) littermate. Typical dorsal white patches Rac1 f/f NRas mouse (arrows). Coat color of forelimb (c) and tail (d) of Rac1 f/f NRas mouse with NRas Ctr littermate. (e) Quantification of white patches on dorsal side of mice. Body weight of mice at P7 (f) and P14 (g); n=6 mice from three different litters. Lower, median, and upper quartile are shown. (h) Ventral skin from P14 NRas Ctr and Rac1 f/f NRas mice with anti-dopachrome tautomerase (melanocytes). Typical cluster of dermal melanocytes in NRas Ctr mouse is shown with an arrow. **P<0.01 compared with Ctr by t-test. Bar=100 μm. NS, not significant.

Mentions: Mice of the C57BL/6 strain normally have light skin and black fur, but when Tyr∷NRasQ61K is expressed, they have black skin and darker fur (Ackermann et al., 2005). Mice lacking Rac1 in melanocytes have white patches on the underside and along the dorsal midline and also a lightening of the paws and tails (Supplementary Figure S2C and D online; Li et al., 2011). At P14, Tyr∷NRasQ61K Rac1 f/f Tyr∷Cre mice had similar pigmentation patterns to Rac1 f/f Tyr∷Cre mice (Figure 2a–d and Supplementary Figure S3A and B online;Li et al., 2011). Furthermore, the number and size of white patches on the dorsal skin between Tyr∷NRasQ61K Rac1 f/f Tyr∷Cre and Rac1 f/f Tyr∷Cre mice was similar (Figure 2a and e and Supplementary Figure S3A online). Mice were born healthy at the expected Mendelian ratio. However, Tyr∷NRasQ61K Rac1 f/f Tyr∷Cre mice were smaller than control littermates and had a similar weight to Rac1 f/f Tyr∷Cre as measured on P7 and P14 (Figure 2f and g and Methods). Histological analysis of P14 Tyr∷NRasQ61K ventral skin revealed melanocytes in hair follicles, dermis and fatty tissue (Figure 2h). Thus, deletion of Rac1 in the melanocyte lineage from mice expressing NRasQ61K resulted in similar pigmentation defects as Rac1 deletion alone.


Activated mutant NRas(Q61K) drives aberrant melanocyte signaling, survival, and invasiveness via a Rac1-dependent mechanism.

Li A, Ma Y, Jin M, Mason S, Mort RL, Blyth K, Larue L, Sansom OJ, Machesky LM - J. Invest. Dermatol. (2012)

Mice expressing melanocyte lineage NRasQ61K retain pigmentation defects caused by Rac1 deletion. Coat color of dorsal (a) and ventral side (b) of P14 Tyr∷NRasQ61K+/o Rac1 f/f Tyr∷Cre+/o (Rac1 f/f NRas) mouse with Tyr∷NRasQ61K+/o (NRas) control (Ctr) littermate. Typical dorsal white patches Rac1 f/f NRas mouse (arrows). Coat color of forelimb (c) and tail (d) of Rac1 f/f NRas mouse with NRas Ctr littermate. (e) Quantification of white patches on dorsal side of mice. Body weight of mice at P7 (f) and P14 (g); n=6 mice from three different litters. Lower, median, and upper quartile are shown. (h) Ventral skin from P14 NRas Ctr and Rac1 f/f NRas mice with anti-dopachrome tautomerase (melanocytes). Typical cluster of dermal melanocytes in NRas Ctr mouse is shown with an arrow. **P<0.01 compared with Ctr by t-test. Bar=100 μm. NS, not significant.
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fig2: Mice expressing melanocyte lineage NRasQ61K retain pigmentation defects caused by Rac1 deletion. Coat color of dorsal (a) and ventral side (b) of P14 Tyr∷NRasQ61K+/o Rac1 f/f Tyr∷Cre+/o (Rac1 f/f NRas) mouse with Tyr∷NRasQ61K+/o (NRas) control (Ctr) littermate. Typical dorsal white patches Rac1 f/f NRas mouse (arrows). Coat color of forelimb (c) and tail (d) of Rac1 f/f NRas mouse with NRas Ctr littermate. (e) Quantification of white patches on dorsal side of mice. Body weight of mice at P7 (f) and P14 (g); n=6 mice from three different litters. Lower, median, and upper quartile are shown. (h) Ventral skin from P14 NRas Ctr and Rac1 f/f NRas mice with anti-dopachrome tautomerase (melanocytes). Typical cluster of dermal melanocytes in NRas Ctr mouse is shown with an arrow. **P<0.01 compared with Ctr by t-test. Bar=100 μm. NS, not significant.
Mentions: Mice of the C57BL/6 strain normally have light skin and black fur, but when Tyr∷NRasQ61K is expressed, they have black skin and darker fur (Ackermann et al., 2005). Mice lacking Rac1 in melanocytes have white patches on the underside and along the dorsal midline and also a lightening of the paws and tails (Supplementary Figure S2C and D online; Li et al., 2011). At P14, Tyr∷NRasQ61K Rac1 f/f Tyr∷Cre mice had similar pigmentation patterns to Rac1 f/f Tyr∷Cre mice (Figure 2a–d and Supplementary Figure S3A and B online;Li et al., 2011). Furthermore, the number and size of white patches on the dorsal skin between Tyr∷NRasQ61K Rac1 f/f Tyr∷Cre and Rac1 f/f Tyr∷Cre mice was similar (Figure 2a and e and Supplementary Figure S3A online). Mice were born healthy at the expected Mendelian ratio. However, Tyr∷NRasQ61K Rac1 f/f Tyr∷Cre mice were smaller than control littermates and had a similar weight to Rac1 f/f Tyr∷Cre as measured on P7 and P14 (Figure 2f and g and Methods). Histological analysis of P14 Tyr∷NRasQ61K ventral skin revealed melanocytes in hair follicles, dermis and fatty tissue (Figure 2h). Thus, deletion of Rac1 in the melanocyte lineage from mice expressing NRasQ61K resulted in similar pigmentation defects as Rac1 deletion alone.

Bottom Line: The Ras oncogene also activates signaling via the related Rho GTPase Rac1, which can mediate growth, survival, and motility signaling.We tested the effects of activated NRas(Q61K) on the proliferation, motility, and invasiveness of melanoblasts and melanocytes in the developing mouse and ex vivo explant culture as well as in a melanoma transplant model.We also show that genetic deletion or pharmacological inhibition of Rac1 in NRas(Q61K) induced melanoma suppresses tumor growth, lymph node spread, and tumor cell invasiveness, suggesting a potential value for Rac1 as a therapeutic target for activated NRas-driven tumor growth and invasiveness.

View Article: PubMed Central - PubMed

Affiliation: The Beatson Institute for Cancer Research, Garscube Estate, Glasgow, UK.

ABSTRACT
Around a fifth of melanomas exhibit an activating mutation in the oncogene NRas that confers constitutive signaling to proliferation and promotes tumor initiation. NRas signals downstream of the major melanocyte tyrosine kinase receptor c-kit and activated NRas results in increased signaling via the extracellular signal-regulated kinase (ERK)/MAPK/ERK kinase/mitogen-activated protein kinase (MAPK) pathways to enhance proliferation. The Ras oncogene also activates signaling via the related Rho GTPase Rac1, which can mediate growth, survival, and motility signaling. We tested the effects of activated NRas(Q61K) on the proliferation, motility, and invasiveness of melanoblasts and melanocytes in the developing mouse and ex vivo explant culture as well as in a melanoma transplant model. We find an important role for Rac1 downstream of NRas(Q61K) in mediating dermal melanocyte survival in vivo in mouse, but surprisingly NRas(Q61K) does not appear to affect melanoblast motility or proliferation during mouse embryogenesis. We also show that genetic deletion or pharmacological inhibition of Rac1 in NRas(Q61K) induced melanoma suppresses tumor growth, lymph node spread, and tumor cell invasiveness, suggesting a potential value for Rac1 as a therapeutic target for activated NRas-driven tumor growth and invasiveness.

Show MeSH
Related in: MedlinePlus