Text

theses

ETD doctoral

The maintenance of apical and basolateral cell polarity in mature epithelial tissue is essential for the tissue homeostasis and function. Cellular polarity is actively maintained by a dedicated group of proteins called the Rab small GTPases, which typically reside in membrane-bound intracellular organelles to engage in the sorting and trafficking of distinct protein and membrane cargoes. Rab11a is a specialized Rab small GTPase that represents an integral component of the recycling endosome. Previous studies in cultured mammalian cells have established Rab11a’s essential role in the formation and maintenance of cellular polarity. However, the in-vivo contribution of Rab11a function to epithelial physiology and pathology remains poorly understood. We have used mouse genetic approach to study the impact of Rab11a loss-of-function on intestinal homeostasis and tumorigenesis. We found that mice with intestinal epithelial loss of Rab11a exhibited early onset of epithelial dysplasia, which synergizes with various risk factors, including aging, chemical carcinogen, and oncogenic pathways to accelerate tumor progression. We found that colorectal cancer tissues with reduced RAB11A displayed poorer survival rates. Mechanistically, transcriptomic analysis of Rab11a-deficient intestinal epithelium uncovered an expansion of the intestinal stem cell signature immediate after Rab11a deletion, and this was accompanied by cell-autonomous activation of Yes-Associated Protein (YAP), the Hippo signaling transducer. Further analysis revealed that human or mouse intestinal epithelial cells lacking RAB11A exhibited diminished YAP phosphorylation, marked nuclear YAP translocation, enhanced IL6-Stat3 and amphiregulin-MAPK signaling activities. Additional cell biology and biochemical investigation demonstrated that RAB11A is critical for YAP to complex with adherens and tight junction components, including α and β-catenin, as well as scaffolding proteins Merlin and AMOTL-2. A novel association between RAB11A and Merlin was identified by us, and Merlin knockdown largely recapitulated RAB11A-deficient changes including YAP activation and cellular hyper-proliferation. Enhanced YAP and β-catenin complexing and nuclear translocation were also demonstrated by us in cells lacking RAB11A. As YAP is physiologically important for epithelial repair, we further demonstrated that mice lacking intestinal epithelial Rab11a were highly susceptible to DSS mediated colonic mucosal injury, and the phenotype likely attributed to an impaired junctional restoration in the injured colon due to abnormal YAP transport to the junction. Our studies suggested that RAB11A-containing recycling endosomes in mature epithelia constitute a key regulatory domain to suppress abnormal growth, restore epithelial junction and barrier integrity after injury via regulating multiple cytokine and growth factor signaling cascades.

ETD_11519

Ph.D.

Includes bibliographical references

doi:10.7282/t3-dm7z-r337