Targeted deletion of the nonmuscle myosin II-B heavy chain (NMHC II-B) in mice caused defects in the heart and brain during embryonic development and homozygous embryos did not survive or those surviving to term died within one day after they were born. Either heart or brain defects might be responsible for the death of homozygous embryos. To further understand the role of NMHC II-B during the development, we are trying to establish transgenic mice specifically overexpressing NMHC II-B in the heart. The transgenic mice are currently being analyzed and will be used in an effort to rescue the knockout mice. The rescued mice will be expected to undergo a normal heart development, but still have defects in the brain. Therefore, the knockout mice rescued by the transgenic mice should allow us to further evaluate the importance of NMHC II-B in the brain. A truncated form of the nonmuscle myosin heavy chain II-A (NMHC II-A), lacking 1-592 amino acids, has also been used to examine its cellular functions. Green fluorescent protein (GFP) was fused to the N-terminal end of both the full-length and truncated NMHC II-A and the fusion proteins were stably expressed in a HeLa Tet-off cell line, in which endogenous NMHC II-A was expressed, but NMHC II-B was not. The expression of truncated NMHC II-A in the HeLa Tet-off cell line induced cell rounding with rearrangements of actin filaments and disappearance of focal adhesions. These changes were reversed when the expression of truncated NMHC II-A was terminated by addition of doxycycline. HeLA Tet-off cells expressing the full-length NMHC II-A:GFP fusion protein did not show this phenotype with or without doxycycline. Confocal microscopic studies indicated that both full-length and truncated NMHC II-A were localized to the stress fibers. We suggest NMHC II-A is involved in maintenance of cell shape by stabilizing the stress fibers and focal adhesions.
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