Mechanisms of Hepatocyte Polarization and Apical Tube Formation

肝细胞极化和心尖管形成的机制

基本信息

批准号：
10221385
负责人：
Erfei Bi
金额：
$ 39.24万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-07-01 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10221385
关键词：
Actin-Binding Protein Actins Adherens Junction Affect Apical Architecture Attenuated Basic Science Bile fluid Binding Biogenesis Biology Blood Circulation Candidate Disease Gene Cell model Cells Cholestasis Cytokinesis Cytoskeleton Defect Development Drug Metabolic Detoxication Epithelial Epithelial Cells Exocytosis Filament Gene Expression Profile Genes Geometry Growth Guanine Nucleotide Exchange Factors Hepatic Cord Hepatocyte Human In Vitro Kinesin Link Liver Liver diseases Location Mediating Membrane Metabolism Microfilaments Microtubules Motor Myosin Type V Natural regeneration Organ Pathway interactions Phosphatidylinositols Play Plus End of the Microtubule Primary carcinoma of the liver cells Process Production Proliferating Public Health Rattus Recycling Role Scaffolding Protein Serum Proteins Side Site Small Interfering RNA Surface Testing Tight Junctions Time Tube Tubular formation Vesicle Visceral base cytohesin-1 daughter cell gene discovery in vivo interdisciplinary approach liver development liver function scaffold spatiotemporal trafficking transcriptomics vesicle transport

Actin-Binding Protein Actins Adherens Junction Affect Apical Architecture Attenuated Basic Science Bile fluid Binding Biogenesis Biology Blood Circulation Candidate Disease Gene Cell model Cells Cholestasis Cytokinesis Cytoskeleton Defect Development Drug Metabolic Detoxication Epithelial Epithelial Cells Exocytosis Filament Gene Expression Profile Genes Geometry Growth Guanine Nucleotide Exchange Factors Hepatic Cord Hepatocyte Human In Vitro Kinesin Link Liver Liver diseases Location Mediating Membrane Metabolism Microfilaments Microtubules Motor Myosin Type V Natural regeneration Organ Pathway interactions Phosphatidylinositols Play Plus End of the Microtubule Primary carcinoma of the liver cells Process Production Proliferating Public Health Rattus Recycling Role Scaffolding Protein Serum Proteins Side Site Small Interfering RNA Surface Testing Tight Junctions Time Tube Tubular formation Vesicle Visceral base cytohesin-1 daughter cell gene discovery in vivo interdisciplinary approach liver development liver function scaffold spatiotemporal trafficking transcriptomics vesicle transport

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项目摘要

Project Summary/Abstract: The functions of the liver, including detoxification, synthesis of serum proteins, and bile production, critically depend on hepatocyte polarization and bile canaliculus (BC) formation. Defects in these processes are associated with serious liver diseases such as cholestasis and hepatocarcinoma. Using the rat hepatocyte line Can 10, the only known cells that can proliferate and form “tubular” BCs (tBCs) in vitro that resembles those in vivo, we discovered that hepatocyte polarization and “primordial” BC formation are linked to cytokinesis. This division-linked mechanism also applies to BC biogenesis during liver development. However, it remains a mystery how a primordial BC formed between two daughter cells at the division site is remodeled and grown into a tBC nestled between two rows of cells, as seen in the liver. We hypothesize that tBC formation involves spatiotemporal coordination of oriented divisions, with BC expansion driven by targeted exocytosis, elongation by pushing forces from both sides of the BC, and extension along the cell-cell contact through constant remodeling of the adherens and tight junctions. To identify the steps and key players involved, we employed two non-biased approaches. The first involves identification of genes from single-cell transcriptomic analysis of liver development that are induced or increased at stages of hepatocyte polarization and BC formation. The second involves identification of genes whose expressions are increased in the polarized Can 10 cells versus the unpolarized parental cells. These complementary approaches have led to the discovery of numerous genes that may play an important role in a process from the terminal stage of cytokinesis to tBC formation. 11 candidate genes were selected for a small siRNA-based screen, three (KIF21B, ABLIM3, and IPCEF1) were found to be required for tBC formation. None of these has been implicated previously in epithelial tube formation. In this application, we will use an interdisciplinary approach to test our hypotheses that the microtubule (MT) plus end-directed motor Kif21B and the actin-binding protein Ablim3 act in concert to drive BC expansion by controlling MT- and actin-mediated vesicle transport as well as adherens junction (AJ) assembly (Aim 1), and that the scaffold protein Ipcef1 promotes Arf6 activation by distinct cytohesins (GEFs) at distinct locations to control AJ remodeling, apical vesicle recycling, and actin-based protrusions to drive BC elongation (Aim 2). As the mechanisms of apical tube formation are highly conserved from worms to humans, the impact of our proposed study will likely reach far beyond the field of liver biology.

项目摘要/摘要：肝脏的功能，包括排毒，血清蛋白的合成以及胆汁产生取决于肝细胞极化和胆管（BC）的形成。这些过程中的缺陷是与严重的肝疾病（例如胆汁淤积和肝癌）有关。使用老鼠肝细胞线CAN 10，唯一可以在体外形成“管状” BCS（TBC）的已知细胞，类似于那些在体内的人，我们发现肝细胞极化和“原始” BC形成与细胞因子。这种分裂连接的机制也适用于肝发育过程中BC生物发生。然而，这仍然是一个神秘的神秘，在分区站点的两个子细胞之间形成原始BC是如何的如肝脏所见，被重塑并生长成一个TBC。我们假设TBC的形成涉及与BC的定向分区的时空协调靶向胞吐作用驱动的扩张，通过从卑诗省两侧推动力的伸长延伸，以及通过粘附和紧密连接的恒定重塑沿细胞接触延伸。到确定所涉及的步骤和主要参与者，我们采用了两种无偏见的方法。第一个涉及通过诱导或在肝细胞极化和BC形成的阶段增加。第二个涉及识别在极化中表达式增加的基因可以10个细胞与非极化亲本细胞相比。这些完整的方法导致发现了许多可能起重要的基因在从细胞因子的末端到TBC形成的过程中的作用。选择了11个候选基因对于基于SIRNA的小屏幕，发现TBC需要三个（KIF21B，ABLIM3和IPCEF1）形成。这些都没有与上皮管形成有关。在此应用程序中，我们将使用跨学科方法来测试我们的假设，即微管（MT）加终端定向 Motor KIF21b和肌动蛋白结合蛋白ABLIM3协同起作用，以控制MT- 和肌动蛋白介导的囊泡运输以及粘附连接（AJ）组件（AIM 1），并且脚手架蛋白IPCEF1在不同位置促进不同位置的不同细胞粘着蛋白（GEF）激活ARF6 AJ重塑，顶囊泡回收和基于肌动蛋白的突起驱动BC延伸（AIM 2）。作为顶端管形成的机制是从蠕虫到人的高度保守的，这是我们的影响拟议的研究可能远远超出了肝生物学领域。