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The role of podocalyxin in adhesion and cell morphogenesis

University of British Columbia

Podocalyxin is a sialomucin expressed on kidney podocytes, vascular endothelia, and hematopoietic progenitors. Although podocalyxin and its close relative, CD34 have been studied for many years, their precise functions have remained elusive, and roles in blocking differentiation, preventing cell adhesion, and establishing cell polarity have all been proposed. Despite this ambiguity, the perinatal lethality of podocalyxin knockout mice (as a result of kidney defects) and podocalyxin’s close association with cancer progression highlight its biological importance. I therefore used several strategies to clarify podocalyxin’s functions and mechanisms of action. Podocalyxin was overexpressed in epithelial cells, and I observed a striking decrease in cell adhesion and an induction of microvillus formation. Microvillus formation was then used as the endpoint to assess the activity of podocalyxin mutants: the extracellular domain was essential while most of the cytoplasmic tail could be deleted without loss of this function. These in vitro studies also demonstrated that podocalyxin recruits the scaffolding protein, NHERF1, which may have important implications in the regulation of NHERF-related processes, such as interaction with ion transporters and signalling molecules. In order to study podocalyxin in vivo, generation of conditional podocalyxin overexpressing mice was attempted. The intention was to generate a single floxed podxl transgenic mouse line that could be crossed with numerous Cre mice in order to induce podocalyxin expression in selected tissues. For example, podocalyxin overexpression in mammary tissue was intended to facilitate evaluation of podocalyxin’s role in breast cancer progression. Similarly, these mice could be used to selectively rescue defects in podocalyxin-deficient mice. Unfortunately, chromosomal abnormalities in the parental embryonic stem cells temporarily prevented completion of this study. As an alternative strategy, we attempted to selectively rescue the kidney defects observed in podocalyxin-null mice by creation of mice with a kidney-specific podxl transgene. Surprisingly, although transgenic podocalyxin was appropriately expressed, and podocyte morphology appeared relatively normal in contrast to podocalyxin-null mice, transgenic mice still died perinatally. This suggests the presence of other serious, as yet undetermined, abnormalities in podocalyxin-deficient animals. Continued assessment of these defects and podocalyxin’s role in cancer progression is underway. In summary, this thesis reveals a new mechanistic role for podocalyxin in the process of cell morphogenesis and suggests that in addition to its vital role in kidney development, podocalyxin may play an essential role in other aspects of mammalian development.

Text

2010-01-18

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http://hdl.handle.net/2429/18619

Medical Genetics

University of British Columbia

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