FUNCTION OF NONMUSCLE MYOSIN II-B HEAVY CHAIN

非肌球蛋白重链-IIB转基因小鼠的建立 （NMHC-IIB）特异性地存在于心肌细胞中，或更普遍地存在于 正在建立，并将与NMHC-IIB缺陷小鼠杂交， 测试将NMHC-IIB加回心肌细胞是否足够 为了防止NMHC-IIB缺陷小鼠产生结构性 心脏缺陷 其结果也将有助于评估效果 NMHC-IIB对非肌肉细胞发育的影响，如 富集NMHC-IIB的神经元细胞。 危险工作据 受NMHC-IIB缺陷小鼠死亡前不久或 出生后 目前，已经构建了NMHC-IIB质粒， 我们的合作者正在生产转基因小鼠。 与人NMHC-IIB cDNA融合的绿色荧光蛋白（GFP）具有 也被用来追踪NMHC-IIB在转染细胞中的定位。 活细胞系 将GFP融合到含有以下的N末端片段： 头颈部结构域分散于细胞质和细胞核中， 转染的COS-7和RBL细胞系，如在GFP对照中所见 构建体 然而，GFP融合到尾部结构域或完整的 人NMHC-IIB cDNA序列定位于这些细胞中，接近 质膜或核周区域。 进一步解剖 尾区显示NMHC-IIB尾区的一个小区域 由约300个氨基酸组成的蛋白质是必需的，并且足以 人NMHC-IIB的定位。

Transgenic mice overexpressing the nonmuscle myosin heavy chain-IIB (NMHC-IIB) specifically in cardiac myocytes or more ubiquitously are being established and will be crossed to the NMHC-IIB deficient mice to test if adding the NMHC-IIB back to the cardiac myocytes is sufficient to prevent the NMHC-IIB deficient mice from developing the structural cardiac defects. The results will also be useful to evaluate effects of the NMHC-IIB on the development of the nonmuscle cells, such as neuronal cells which are enriched in NMHC-IIB. The latter has been plagued by the fact that NMHC-IIB deficient mice died shortly before or after birth. Currently, the NMHC-IIB plasmid has been constructed and the transgenic mice are being produced with our collaborators. The green fluorescent protein (GFP) fused to human NMHC-IIB cDNA has also been used to trace the localization of the NMHC-IIB in transfected living cell lines. The GFP fused to a N-terminal fragment containing the head and neck domain dispersed in the cytoplasm and nucleus of the transfected COS-7 and RBL cell lines, as was also seen with GFP control construct. However, the GFP fused to the tail domain or the complete sequence of human NMHC-IIB cDNA was localized in these cells, close to the plasma membrane, or to perinuclear regions. Further dissection of the tail region showed that a small domain of the NMHC-IIB tail comprised of about 300 amino acids is necessary and sufficient for the localization of the human NMHC-IIB.