Using Reflection Interference Contrast Microscopy to Differentiate Focal Adhesions from Passive Adhesions in Adherent Cells

Russel, Jasmin G. (2015) Using Reflection Interference Contrast Microscopy to Differentiate Focal Adhesions from Passive Adhesions in Adherent Cells. Masters thesis, Indian Institute of Science Education and Research Kolkata.

PDF (MS dissertation of Jasmin G. Russel (10MS68)) Jasmin_G_Russel_10MS68_MS_Thesis.pdf - Submitted Version Restricted to Repository staff only Download (1MB)

Abstract

Focal adhesions (FAs) are specialized structures that connect the actin cytoskeleton of a eukaryotic cell to the extracellular matrix via cell membrane through integrin family of receptors – and hence form “regulated” sites where force transduction occurs from the inside of the cell to its surrounding and vice versa. Force transduction to the membrane merely needs regions of adhesion, while the presence of filamentous actin is necessary to transduce the force to other force-bearing structures of the cell. Passive adhesions are mere electrostatic attraction of cell membrane and substrate – and, we hypothesize, can well forms regions of force transduction albeit transient and with lower depth of transduction. In my MS thesis I quantify the relative presence of focal vs passive adhesion regions in live cells (HeLa) and to compare their lifetimes over long timescales in the same cell. Using Reflection Interference Contrast Microscopy (RICM), regions of close contacts (appear as dark patches) are found out. Comparing RICM images with Pax-GFP (Paxillin – focal adhesion marker) imaging on the same cell, we “learn” how to identify focal adhesions and what percentage regions are passive adhesions. The focal adhesions play a major role in determining the fate of a cell, signal transduction, cell motility etc. They are very much important for the survival and integrity of cellular activities. My thesis will set an assay to identify FAs noninvasively (without transfection) and also throw new light on the net contribution of passive adhesions in force transduction.

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