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Technical Data Sheet

MK7730

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Myosin Light Chain Phospho-Regulation
Immunocytochemistry Kit
Price

$455

Kit Components
Applications: WB = Western blot, E = ELISA, ICC = Immunocytochemistry, IP = Immunoprecipitation, IHC = Immunohistochemistry Species: H = Human, R = Rat, M = Mouse, C = Chicken, F = Fish

Product References
MP4221 Ho, H. et al. (2013) Pigment Cell Melanoma Res 26(2):218. WB: human A375 melanoma
MP3461 Aguilar, H.N. et al. (2011) PLoS ONE 6(6):e20903. WB: human uterine myocytes
MP3461 Beach, J.R. et al. (2011) BMC Cell Biol. 12:52. ICC: PHK and HeLa cells
Kit Summary
The Myosin Regulatory Light Chain phospho-regulation kit can be used for immunocytochemical co-localization of MLC phosphorylation at Ser-1 and Ser-19 compared to the total expression of MLC. The kit includes rabbit polyclonal and mouse monoclonal antibodies along with Goat-anti-rabbit conjugated to DyLight® 594 and Goat anti-mouse conjugated to DyLight® 488 for dual labeling experiments.
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Enlarge

Immunocytochemical labeling of phosphorylated MLC in paraformaldehyde fixed A7r5 cells. The cells were dual-labeled with anti-MLC (MM3441; middle) and anti-MLC (MP4201; top left), anti-MLC (Ser-19) (MP4221; middle left) and anti-MLC (Ser-1) (MP3461; bottom left). Goat anti-Mouse DyLight® 488 and Goat anti-Rabbit DyLight® 594 were used for detection of primary antibodies. The overlay of staining patterns are shown to the right.

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Background
Both smooth muscle and non-muscle myosin II activity is regulated by the phosphorylation state of the myosin regulatory light chain (MLC, MRLC, MLC20, Myl9). Phosphorylation of MLC at Thr-18 and Ser-19 activates myosin II motor activity and increases myosin filament stability. This activation has important roles in various cell motile processes. By contrast, other phosphorylation sites on MLC may inhibit myosin II activity. PKC phosphorylates Ser-1/Ser-2 and Thr-9 in MLC, and this phosphorylation decreases activated myosin II interaction with actin, as well as inhibits MLC interaction with the activation site kinase, myosin light-chain kinase. The Ser-1/Ser-2 region may be the major inhibitory site since Ser-1 is phosphorylated during PDGF-induced stress fiber disassembly and expression of unphosphorylatable MLC20 at the Ser-1/Ser-2 site suppresses this disassembly. Thus, inhibition of myosin II activity through phosphorylation of Ser-1/Ser-2 may have important roles in growth factor-induced reorganization of actomyosin filaments.

Background References
Sellers, J.R. (1991) Curr. Opin. Cell Biol. 3:98.
Tan, J.L. et al. (1992) Annu. Rev. Biochem. 61:721.
Komatsu, S. & Ikebe, M. (2007) Mol. Biol. Cell 18:5081.
Buffer and Storage
Mouse monoclonal, rabbit polyclonal, and secondary reagents are supplied in phosphate-buffered saline, 50% glycerol, 1 mg/ml BSA, and 0.05% sodium azide. Store at –20°C. Stable for 1 year.
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