Mayanil Laboratory Main

Mayanil Laboratory

Projects:

1. Neural Crest stem cells and the Gene Regulatory Network

Pax3 regulates neural crest cell migration and is critical during neural crest development. TGFβs modify neural crest cell migration and differentiation. TGFβ2 nullizygous embryos (TGFβ2^-/-Pax3^+/+) display open neural tube and bifid spine, whereas in wild type embryos the neural tube is closed. We have demonstrated that Pax3 regulates TGFβ2 by directly binding to cis-regulatory elements on its promoter. Later we found that the TGFβ2 nullizygous phenotype can be reversed to the wild type phenotype by down-regulating one allele of Pax3, as in TGFβ2^-/-Pax3^+/-embryos obtained through breeding TGFβ2^+/-Pax3^+/- mice. Thus Pax3 and TGFβ2 interact in a coordinated gene regulatory network, linked by common downstream effector genes, to bring about this phenotypic reversal. Downstream effectors include Hes1, Ngn2 and Sox9, as well as other genes involved in neuronal differentiation are being currently investigated.

2. Neurogenesis permissive chromatin during caudal neural tube development

The central question in embryonic development is how neural stem cells switch from being proliferative to getting differentiated. During neurogenesis stem cells differentiate into neurons, which no longer proliferate but extend nerve fibers. Although several molecules are known to inhibit cell proliferation during neuronal differentiation, the pathways of cell-division suppression are not fully identified. Our lab has recently shown that Pax3 regulates Hes1 and Ngn2 promoters by directly interacting with the cis-regulatory regions within their promoters, nothing is known about the chromatin structures at these candidate Pax3 target promoters that lead to neurogenesis during caudal neural tube development. Our lab is focused on trying to identify “Neurogenesis permissive chromatin” by the analyzing distinct pattern of differently modified histones during neurogenesis.

3. Mechanisms of Epigenetic Rescue of Neural Tube closure defects by Folic Acid

FA rescue

Hypothetical model for FA mediated rescue of stem cell proliferation and neurogenesis. Pax3 mutant “Splotch” homozygous embryos do not express functional Pax3 which leads to up-regulation of several miRNAs, some of which target H3K27 histone demethylase KDM6B. This results in decreased KDM6B expression and increased H3K27 methylation. The association between methylated H3K27 and the Hes1 promoter results in (a) decreased Hes1 expression and lowered stem cell proliferation. At the same time low association between methylated H3K27 and Neurog2 promoter, but high association between acetylated H3K9 and H3K18 and Neurog2 promoter results in (b) premature neurogenesis. FA mediated rescue could be due to reversible down-regulation of KDM6B targeting microRNAs, resulting in up-regulation of KDM6B and subsequent lowering of H3K27 methylation. Demethylated H3K27 shows decreased binding to Hes1 promoter and increased binding to Neurog2 promoter, which results in (c) increased stem cell proliferation/maintenance and (d) normal neurogenesis.

4. Discovery and Validation of Epigenetic markers for Spina Bifida

Based on the model described above, we hypothesized that the Spina Bifida fetus would shed out stem cells from the open neural tube portion into the amniotic fluid. These stem cells in the amniotic fluid of the woman carrying Spina Bifida fetus would show higher expression of H3K27me2 mark. The above data confirms the hypothesis. The stem cell marker, Oct4 is very high in amniotic fluid of woman carrying Spina Bifida fetus than a normal fetus. The H3K27 methylation mark is also higher as expected.

Publications (h index = 11)

Nakazaki, H., Shen, Y-W., Yun, B., Reddy, A. C., Khanna, V., Ichi, S., Mania-Farnell, B., McLone, D. G. and Tomita, T. and Mayanil, C. S. (2009) Transcriptional Regulation by Pax3 and TGFβ2 signaling constitute a gene regulatory network. Int. J. Dev. Biol. 53: 69-79.
Nakazaki H., Reddy, A. C., Mania-Farnell B., McCabe, C., Ichi, S., George, D., McLone D. G., Tomita T. and Mayanil, C. S. (2008) Key basic helix loop helix transcription factor genes Hes1 and Ngn2 are regulated by Pax3 during mouse embryonic development. Dev. Biol. 316: 510-523.
Mayanil, C. S., Pool, A., Nakazaki, H., Reddy, A., Mania-Farnell, B., Yun, B., George, D., McLone, D. G., and Bremer, E. G. (2006) Regulation of murine TGFβ2 by Pax3 during early embryonic development. J. Biol. Chem. 281: 24544-24552.
Knepper, P. A., Mayanil, C.S., Goossens, W., Wertz, R.D., Holgren, C., Ritch, R., and Allingham, R.R. (2002) Aqueous humor in primary open-angle glaucoma contains an increased level of CD44S. Invest Ophthalmol. Vis. Sci. 43(1):133-139.
Mayanil, C.S., George, D., Freilich, L., Miljan, E.J., Mania-Farnell, B., McLone, D.G. and Bremer, E.G. (2001) Microarray Analysis Detects Novel Pax3 Downstream Target Genes. J. Biol. Chem. 276, 49299-49309.
Mayanil, C.S., George, D., Mania-Farnell, B., Bremer, C., McLone, D. G. and Bremer, E. G. (2000) Over-expression of murine Pax3 increases NCAM polysialylation in a human medulloblastoma cell line. J. Biol. Chem. 275: 23259-23266.