Discovery and function of ion channels in liver epithelia.

我的研究重点是离子通道在肝脏生理学中的识别和作用。我们目前的重点是瞬时受体潜能Melastatin(TRPM)通道，异构体7。TRPM7是一种普遍表达的通道激酶，与许多增殖细胞的存活有关；敲除是胚胎致命的。最近的研究支持TRPM7在器官发生中的重要作用。我们发现在分裂的肝细胞中TRPM7通道的活性显著高于终末分化的、未分裂的成年肝细胞。与成年细胞相比，通道阻断导致分裂时的死亡率更高，这表明镁离子内流在支持肝细胞增殖中起着基础性作用。我们还将TRPM7定位于所有肝细胞的核膜，除了在其他细胞中所见的胞浆点状表达外。TRPM7在未分裂的成年肝细胞核膜中的表达高于分裂细胞。分化条件降低了分裂细胞的通道活性，增加了核膜TRPM7。在有丝分裂过程中，TRPM7与有丝分裂纺锤体相联系。因此，TRPM7通道活性似乎在胚胎肝细胞扩张和肝癌细胞存活中起重要作用，而核膜被膜的积累与终末分化有关。在接下来的五年里，我们将确定TRPM7如何支持胚胎肝细胞的扩张和去分化生长，以及终末分化。将使用从分裂的胚胎细胞和肝癌细胞到未分裂的成年细胞的大鼠肝细胞的光谱。这些细胞为研究TRPM 7在同一组织和物种的分裂和非分裂细胞中的表达和功能以及研究其在肝脏发育中的作用提供了独特的机会。基因干扰和过度表达技术以及通道突变结构将被用来补充电生理和分子方法，以确定将TRPM7的表达、定位和活性与胚胎扩张、非调控增殖和终末分化联系起来的分子事件。四个目标被描述，为5-6个研究生论文和许多本科项目提供基础。

My research program is focused on the identification and roles of ion channels in the physiology of the liver. Our current focus is on a Transient Receptor Potential Melastatin (TRPM) channel, isoform 7. TRPM7, a ubiquitously expressed channel-kinase, is associated with survival of many proliferating cells; knock-out is embryonic lethal. Recent studies support a significant role for TRPM7 in organogenesis. We showed that TRPM7 channel activity is significantly higher in dividing liver cells than in terminally differentiated, non-dividing adult hepatocytes. Channel block caused higher death rates in dividing as compared with adult cells, implying a fundamental role for Mg2+ influx in supporting liver cell proliferation. We also localized TRPM7 to the nuclear envelope in all liver cells in addition to the established cytosolic, punctate expression seen in other cells. TRPM7 expression was higher in the nuclear envelope of non-dividing adult hepatocytes as compared with dividing cells. Differentiating conditions reduced channel activity and increased nuclear envelope TRPM7 in dividing cells. During mitosis TRPM7 associates with the mitotic spindle. Thus TRPM7 channel activity seems to be important in embryonic hepatocyte expansion and hepatoma cell survival, whereas nuclear envelope accumulation is associated with terminal differentiation. Over the next five years we will define how TRPM7 supports embryonic hepatocyte expansion and de-differentiated growth, and terminal differentiation. A spectrum of rat hepatocytes from dividing embryonic and hepatoma cells to non-dividing adult cells will be used. These cells provide unique opportunities to study TRPM 7 expression and function in dividing and non-dividing cells from the same tissue and species and to study its role in liver development. Gene disruption and over-expression techniques and channel mutant constructs will be used to complement electrophysiological and molecular approaches to identify the molecular events linking TRPM7 expression, localization and activity to embryonic expansion, unregulated proliferation and terminal differentiation. Four aims are described providing the basis for 5-6 graduate theses and numerous undergraduate projects.