: Emerging data suggest that impaired motility of the colon could be associated with the absence of phosphorylation of the small heat shock protein HSP27: (1) Transgenic mice over-expressing non-phospho-HSP27 showed inhibition of acetylcholine-induced contraction (decrease in cell length), (2) Smooth muscle cells from the colons of aged rats, exhibited 50% decreased contraction with concomitant decreased phosphorylation of HSP27 and decreased association of actin with myosin. HSP27 phosphorylation seems to be at the core of the PKC-mediated sliding of filaments in smooth muscle cells of the colon, by modulating the interaction of the thin filament proteins (tropomyosin, caldesmon and calponin): (1) association of tropomyosin with HSP27 is modulated by phosphorylation of HSP27;(2) non-phosphomimic HSP27 transfected colonic smooth muscle show: (a) reduced association of tropomyosin with HSP27, (b) reduced association of actin with myosin, (c) decreased phosphorylation of caldesmon, and (d) inhibition of the decreased association of tropomyosin with phospho-caldesmon. Our preliminary data also indicate that we were able to reinstate the association of HSP27 with tropomyosin in non-phosphomimic HSP27 transfected colonic smooth muscle cells by introducing phosphomimic HSP27. Specifically we propose to study the role of PKC-mediated phosphorylated HSP27, in modulating the activities of thin filament regulatory proteins (leading to contraction of circular colon smooth muscle). The coordinated phosphorylation and interactions of thin filament regulatory proteins with each other, with actin as well as with HSP27 may modulate thin filament based regulation of contraction in circular smooth muscle cells of colon. We therefore propose to: (1) Examine PKC-mediated phosphorylation of tropomyosin, caldesmon, and calponin. (2) Examine the effect of PKC-mediated phosphorylated HSP27 on the phosphorylation of caldesmon, calponin, and tropomyosin. Investigate the role of phosphorylated HSP27 in coordinated phosphorylation and interactions of thin filament regulatory proteins with each other and with actin. Investigate the effect of phosphorylated HSP27 on their individual and coordinated role in actomyosin interaction. And (3) Use the phospho/non-phospho-HSP27 mutant transfected circular colonic smooth muscle cells and smooth muscle cells from the colons of phospho/non-phospho-HSP27 mutant transgenic mice, to examine the effect of phosphorylated HSP27 on: (a) the PKC-mediated phosphorylation of tropomyosin, caldesmon and calponin, (b) their interaction, (c) modulation of myosin light chain phosphorylation.
| Raghavan, Shreya; Lam, Mai T; Foster, Lesley L et al. (2010) Bioengineered three-dimensional physiological model of colonic longitudinal smooth muscle in vitro. Tissue Eng Part C Methods 16:999-1009 |
| Somara, Sita; Gilmont, Robert R; Varadarajan, Saranyaraajan et al. (2010) Phosphorylated HSP20 modulates the association of thin-filament binding proteins: caldesmon with tropomyosin in colonic smooth muscle. Am J Physiol Gastrointest Liver Physiol 299:G1164-76 |
| Somara, Sita; Gilmont, Robert; Bitar, Khalil N (2009) Role of thin-filament regulatory proteins in relaxation of colonic smooth muscle contraction. Am J Physiol Gastrointest Liver Physiol 297:G958-66 |
| Somara, Sita; Bitar, Khalil N (2008) Direct association of calponin with specific domains of PKC-alpha. Am J Physiol Gastrointest Liver Physiol 295:G1246-54 |
| Somara, Sita; Bitar, Khalil N (2006) Phosphorylated HSP27 modulates the association of phosphorylated caldesmon with tropomyosin in colonic smooth muscle. Am J Physiol Gastrointest Liver Physiol 291:G630-9 |
| Goldsmith, Adam M; Bentley, J Kelley; Zhou, Limei et al. (2006) Transforming growth factor-beta induces airway smooth muscle hypertrophy. Am J Respir Cell Mol Biol 34:247-54 |
| Bitar, Khalil N (2005) Aging and Gi smooth muscle fecal incontinence: Is bioengineering an option. Exp Gerontol 40:643-9 |
| Somara, Sita; Pang, Haiyan; Bitar, Khalil N (2005) Agonist-induced association of tropomyosin with protein kinase Calpha in colonic smooth muscle. Am J Physiol Gastrointest Liver Physiol 288:G268-76 |
| Patil, Suresh B; Pawar, Mercy D; Bitar, Khalil N (2004) Direct association and translocation of PKC-alpha with calponin. Am J Physiol Gastrointest Liver Physiol 286:G954-63 |
| Somara, Sita; Bitar, Khalil N (2004) Tropomyosin interacts with phosphorylated HSP27 in agonist-induced contraction of smooth muscle. Am J Physiol Cell Physiol 286:C1290-301 |
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