Evidence for Single Subunit Surface Expression of Drosophila Integrins and a Possible, Unconventional Method of Transport

Date of Award

2010

Document Type

Thesis

Degree Name

Bachelors

Department

Natural Sciences

First Advisor

Clore, Amy

Keywords

Integrin, Secretory Pathway, Unconventional Transport, Surface Proteins

Area of Concentration

Biology

Abstract

The conventional secretory pathway describes the transport of certain proteins (e.g., those destined to be secreted or displayed in the plasma membrane) from the site of their nascent synthesis to their final location within the cell. According to this pathway, once a protein is completely assembled and properly folded in the endoplasmic reticulum (ER) it is transferred via vesicles to the Golgi apparatus, where it undergoes further modification, and in the case of membrane proteins, is eventually transported in vesicles to the cell surface. Integrins are heterodimeric transmembrane cell surface proteins each composed of α and β subunits. They play an important role in both bidirectional cell signaling and cell migration and adhesion. Integrins are responsible for epithelial cell movement during development, leukocyte �walking� towards a region of inflammation, platelet movement to an infection site, tumor metastasis, and other functions. Until recently, integrins were typically believed to follow the conventional secretory pathway, in which their α and β subunits would assemble in the ER before being transported to the Golgi. Failure to assemble has traditionally been thought to result in retention in the ER, eventual degradation by the ERquality control system, and the inability to reach the cell surface. Recently, however, Schotman et al. (2008) discovered that the Drosophila melanogaster αPS1 integrin subunit, but not the βPS subunit, could reach the plasma membrane in a Golgi-independent manner in Drosophila embryos during a specific stage of development. Previously unexplored data (Bunch and Brower, 1992) and unpublished observations (T.A. Bunch) implied that the βPS subunit might be expressed on the cell surface independently of the αPS2 subunit in a cell culture system. Collectively, the data suggest that the integrin subunits can utilize different, perhaps unconventional, modes of transport and be expressed independently on the cell surface. Unconventional transport models are thought to occur in other systems as well and are of interest as both novel biological pathways, and as possible targets for pharmaceuticals. In the present work, we studied the surface expression of the αPS2 and βPS integrin subunits in transformed Drosophila cells using flow cytometry and immunoblot analysis, to determine if these subunits could be expressed independently on the cell surface. We also explored the effect of using RNAi to downregulate certain genes important in protein assembly and in conventional secretory transport to determine if they had an effect on the surface expression of integrin subunits that might go by an alternative pathway. Taken together, our results suggest that the βPS subunit is able to reach the cell surface independently in cultured Drosophila cells by a route that is not dependent on dGRASP, Syntaxin-5, and calnexin and calreticulin in the same manner as the conventional secretory pathway.

Rights

This bibliographic record is available under the Creative Commons CC0 public domain dedication. The New College of Florida, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.

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