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Technical Data Sheet

MK6490

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Myosin Light Chain Phospho-Regulation
Antibody Sampler Kit
Price

$395

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
RS3251 Kawasaki, H. et al. (2013) World J Gastroenter. 19(17):2629. WB, ICC: mouse intestinal myofibroblasts
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
MS3001 Estrada-Bernal, A. et al. (2011) J Neurooncol. 102:353. Western Blot
RS3251 Estrada-Bernal, A. et al. (2011) J Neurooncol. 102:353. Western Blot
Kit Summary
The myosin regulatory light chain (MLC) phospho-regulation antibody sampler kit can be used to examine phosphorylation of MLC at Ser-1 and Ser-19. The kit also includes monoclonal and polyclonal antibodies to monitor total expression levels of MLC, as well as secondary reagents for antibody detection.
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Western blot analysis of C2C12 cells untreated (lanes 1, 3, 5, & 7) or treated with Lambda phosphatase (lanes 2, 4, 6, & 8). The blots were probed with monoclonal anti-MLC20 (clone MY-21) (lanes 1 & 2), polyclonal anti-MLC (Ser-19) phosho-specific (lanes 3 & 4), anti-MLC (Ser-1) phosho-specific (lanes 5 & 6), or anti-MLC (a.a. 11-22) (lanes 7 & 8).

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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 nonmuscle 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 and rabbit polyclonal antibodies are supplied in phosphate-buffered saline, 50% glycerol, 1 mg/ml BSA, and 0.05% sodium azide. The secondary reagents are supplied in the same buffer without azide. Store all at –20°C. Stable for 1 year.
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