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Transmembrane potential of GlyCl-expressing instructor cells induces a neoplastic-like conversion of melanocytes via a serotonergic pathway.

Blackiston D, Adams DS, Lemire JM, Lobikin M, Levin M - Dis Model Mech (2010)

Bottom Line: Molecular-genetic depolarization of a sparse, widely distributed set of GlyCl-expressing cells non-cell-autonomously induces a neoplastic-like phenotype in melanocytes: they overproliferate, acquire an arborized cell shape and migrate inappropriately, colonizing numerous tissues in a metalloprotease-dependent fashion.A similar effect was observed in human melanocytes in culture.Depolarization of GlyCl-expressing cells induces these drastic changes in melanocyte behavior via a serotonin-transporter-dependent increase of extracellular serotonin (5-HT).

View Article: PubMed Central - PubMed

Affiliation: Center for Regenerative and Developmental Biology, Biology Department, 200 Boston Avenue, Suite 4600, Tufts University, Medford, MA 02155, USA.

ABSTRACT
Understanding the mechanisms that coordinate stem cell behavior within the host is a high priority for developmental biology, regenerative medicine and oncology. Endogenous ion currents and voltage gradients function alongside biochemical cues during pattern formation and tumor suppression, but it is not known whether bioelectrical signals are involved in the control of stem cell progeny in vivo. We studied Xenopus laevis neural crest, an embryonic stem cell population that gives rise to many cell types, including melanocytes, and contributes to the morphogenesis of the face, heart and other complex structures. To investigate how depolarization of transmembrane potential of cells in the neural crest's environment influences its function in vivo, we manipulated the activity of the native glycine receptor chloride channel (GlyCl). Molecular-genetic depolarization of a sparse, widely distributed set of GlyCl-expressing cells non-cell-autonomously induces a neoplastic-like phenotype in melanocytes: they overproliferate, acquire an arborized cell shape and migrate inappropriately, colonizing numerous tissues in a metalloprotease-dependent fashion. A similar effect was observed in human melanocytes in culture. Depolarization of GlyCl-expressing cells induces these drastic changes in melanocyte behavior via a serotonin-transporter-dependent increase of extracellular serotonin (5-HT). These data reveal GlyCl as a molecular marker of a sparse and heretofore unknown cell population with the ability to specifically instruct neural crest derivatives, suggest transmembrane potential as a tractable signaling modality by which somatic cells can control stem cell behavior at considerable distance, identify a new biophysical aspect of the environment that confers a neoplastic-like phenotype upon stem cell progeny, reveal a pre-neural role for serotonin and its transporter, and suggest a novel strategy for manipulating stem cell behavior.

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Hyperpigmentation is due to depolarization. Microinjection of a dominant-negative form of ductin (dn-xDuct) at the one-cell stage inhibits the hyperpolarizing H+-V-ATPase and results in hyperpigmentation (A,B). Injections result in hyperpigmented tadpoles in 11.5% of embryos (C), significantly higher than background levels observed in control embryos. Hyperpigmented embryos arising from dn-xDuct injections were photographed and the number of melanocytes in the eye field counted; there was a 2.1-fold increase in number of melanocytes compared with age-matched controls (D). By contrast, overexpression of the hyperpolarizing potassium channel Kir4.1 (E,F) inhibits ivermectin-induced hyperpigmentation in 25% of injected embryos (G). Kir4.1-mediated inhibition was non-cell-autonomous, because one of two cell injections, resulting in hyperpolarizing channel activity on just one side of the embryo, inhibited hyperpigmentation on both the left and right side of the embryos (H).
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f5-0040067: Hyperpigmentation is due to depolarization. Microinjection of a dominant-negative form of ductin (dn-xDuct) at the one-cell stage inhibits the hyperpolarizing H+-V-ATPase and results in hyperpigmentation (A,B). Injections result in hyperpigmented tadpoles in 11.5% of embryos (C), significantly higher than background levels observed in control embryos. Hyperpigmented embryos arising from dn-xDuct injections were photographed and the number of melanocytes in the eye field counted; there was a 2.1-fold increase in number of melanocytes compared with age-matched controls (D). By contrast, overexpression of the hyperpolarizing potassium channel Kir4.1 (E,F) inhibits ivermectin-induced hyperpigmentation in 25% of injected embryos (G). Kir4.1-mediated inhibition was non-cell-autonomous, because one of two cell injections, resulting in hyperpolarizing channel activity on just one side of the embryo, inhibited hyperpigmentation on both the left and right side of the embryos (H).

Mentions: If hyperpigmentation is truly a result of membrane depolarization, then cellular voltage modulators that function independently of chloride channels should result in the same phenotype. We therefore tested the effect of disrupting the H+-V-ATPase hyperpolarizing pump, which we previously showed plays an important role in several voltage-regulated events in Xenopus development (Adams et al., 2007; Adams et al., 2006) and controls cellular Vmem through movement of H+, not Cl−, ions. Injection of low levels of mRNA encoding a well-characterized dominant-negative mutant of Ductin (the 16-kDa proteolipid subunit c of the hyperpolarizing V-ATPase pump complex) into one-cell embryos resulted in broad, long-lasting (tracked by fused YFP; data not shown) expression, which induced hyperpigmentation in 11.5% of embryos, significantly higher than background hyperpigmentation observed in controls (binomial calculation; n=189 for controls, n=192 for DN-Ductin, P≤0.001) (Fig. 5A–C). Moreover, dn-xDuct-injected hyperpigmented embryos had 2.1-fold more melanocytes than control animals at stage 46 (Student’s t-test, t=7.37, n=11 per treatment, P≤0.001; Fig. 5D).


Transmembrane potential of GlyCl-expressing instructor cells induces a neoplastic-like conversion of melanocytes via a serotonergic pathway.

Blackiston D, Adams DS, Lemire JM, Lobikin M, Levin M - Dis Model Mech (2010)

Hyperpigmentation is due to depolarization. Microinjection of a dominant-negative form of ductin (dn-xDuct) at the one-cell stage inhibits the hyperpolarizing H+-V-ATPase and results in hyperpigmentation (A,B). Injections result in hyperpigmented tadpoles in 11.5% of embryos (C), significantly higher than background levels observed in control embryos. Hyperpigmented embryos arising from dn-xDuct injections were photographed and the number of melanocytes in the eye field counted; there was a 2.1-fold increase in number of melanocytes compared with age-matched controls (D). By contrast, overexpression of the hyperpolarizing potassium channel Kir4.1 (E,F) inhibits ivermectin-induced hyperpigmentation in 25% of injected embryos (G). Kir4.1-mediated inhibition was non-cell-autonomous, because one of two cell injections, resulting in hyperpolarizing channel activity on just one side of the embryo, inhibited hyperpigmentation on both the left and right side of the embryos (H).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3008964&req=5

f5-0040067: Hyperpigmentation is due to depolarization. Microinjection of a dominant-negative form of ductin (dn-xDuct) at the one-cell stage inhibits the hyperpolarizing H+-V-ATPase and results in hyperpigmentation (A,B). Injections result in hyperpigmented tadpoles in 11.5% of embryos (C), significantly higher than background levels observed in control embryos. Hyperpigmented embryos arising from dn-xDuct injections were photographed and the number of melanocytes in the eye field counted; there was a 2.1-fold increase in number of melanocytes compared with age-matched controls (D). By contrast, overexpression of the hyperpolarizing potassium channel Kir4.1 (E,F) inhibits ivermectin-induced hyperpigmentation in 25% of injected embryos (G). Kir4.1-mediated inhibition was non-cell-autonomous, because one of two cell injections, resulting in hyperpolarizing channel activity on just one side of the embryo, inhibited hyperpigmentation on both the left and right side of the embryos (H).
Mentions: If hyperpigmentation is truly a result of membrane depolarization, then cellular voltage modulators that function independently of chloride channels should result in the same phenotype. We therefore tested the effect of disrupting the H+-V-ATPase hyperpolarizing pump, which we previously showed plays an important role in several voltage-regulated events in Xenopus development (Adams et al., 2007; Adams et al., 2006) and controls cellular Vmem through movement of H+, not Cl−, ions. Injection of low levels of mRNA encoding a well-characterized dominant-negative mutant of Ductin (the 16-kDa proteolipid subunit c of the hyperpolarizing V-ATPase pump complex) into one-cell embryos resulted in broad, long-lasting (tracked by fused YFP; data not shown) expression, which induced hyperpigmentation in 11.5% of embryos, significantly higher than background hyperpigmentation observed in controls (binomial calculation; n=189 for controls, n=192 for DN-Ductin, P≤0.001) (Fig. 5A–C). Moreover, dn-xDuct-injected hyperpigmented embryos had 2.1-fold more melanocytes than control animals at stage 46 (Student’s t-test, t=7.37, n=11 per treatment, P≤0.001; Fig. 5D).

Bottom Line: Molecular-genetic depolarization of a sparse, widely distributed set of GlyCl-expressing cells non-cell-autonomously induces a neoplastic-like phenotype in melanocytes: they overproliferate, acquire an arborized cell shape and migrate inappropriately, colonizing numerous tissues in a metalloprotease-dependent fashion.A similar effect was observed in human melanocytes in culture.Depolarization of GlyCl-expressing cells induces these drastic changes in melanocyte behavior via a serotonin-transporter-dependent increase of extracellular serotonin (5-HT).

View Article: PubMed Central - PubMed

Affiliation: Center for Regenerative and Developmental Biology, Biology Department, 200 Boston Avenue, Suite 4600, Tufts University, Medford, MA 02155, USA.

ABSTRACT
Understanding the mechanisms that coordinate stem cell behavior within the host is a high priority for developmental biology, regenerative medicine and oncology. Endogenous ion currents and voltage gradients function alongside biochemical cues during pattern formation and tumor suppression, but it is not known whether bioelectrical signals are involved in the control of stem cell progeny in vivo. We studied Xenopus laevis neural crest, an embryonic stem cell population that gives rise to many cell types, including melanocytes, and contributes to the morphogenesis of the face, heart and other complex structures. To investigate how depolarization of transmembrane potential of cells in the neural crest's environment influences its function in vivo, we manipulated the activity of the native glycine receptor chloride channel (GlyCl). Molecular-genetic depolarization of a sparse, widely distributed set of GlyCl-expressing cells non-cell-autonomously induces a neoplastic-like phenotype in melanocytes: they overproliferate, acquire an arborized cell shape and migrate inappropriately, colonizing numerous tissues in a metalloprotease-dependent fashion. A similar effect was observed in human melanocytes in culture. Depolarization of GlyCl-expressing cells induces these drastic changes in melanocyte behavior via a serotonin-transporter-dependent increase of extracellular serotonin (5-HT). These data reveal GlyCl as a molecular marker of a sparse and heretofore unknown cell population with the ability to specifically instruct neural crest derivatives, suggest transmembrane potential as a tractable signaling modality by which somatic cells can control stem cell behavior at considerable distance, identify a new biophysical aspect of the environment that confers a neoplastic-like phenotype upon stem cell progeny, reveal a pre-neural role for serotonin and its transporter, and suggest a novel strategy for manipulating stem cell behavior.

Show MeSH
Related in: MedlinePlus