Determining the Fate of NKCC1 in the Endocytic Pathway and its Role in Host Defense

Presentation Type

Poster Presentation

Symposium Date

Summer 7-30-2018

Abstract

Fluid secretion represents an important defense mechanism of the gut. Driven by transepithelial chloride section, it flushes pathogens from the intestines. The basolateral Na-K-2Cl cotransporter 1 (NKCC1) is the main mechanism for loading the cell with chloride. Activation of the protein kinase C (PKC) causes internalization of NKCC1 and consequently, a decrease in fluid secretion. However, it is unknown whether the internalization pathway is one of degradation or recycling. In the colon, opportunistic pathogens, such as the fungus Candida albicans (C. albicans) invade the body by shifting its morphology from yeast to filamentous. We hypothesize that when pathogenic, C.albicans decreases fluid secretion by causing NKCC1 internalization.

Our research seeks to (1) determine the fate of NKCC1 in the internalization pathway; and (2) explore how C. albicans, when made pathogenic (i.e., filamentous), interferes with the intestine’s secretory defenses.

Previously, the lab has attempted to map NKCC1 in the endocytic pathway using microscopy and endosomal markers. The results suggested that NKCC1 may recycle to the plasma membrane. Since then, we have adopted a biochemical approach using western blotting with T84 and MDCK cells. Our experiments blocked possible degradation pathways (lysosome and proteasome), and the amount of NKCC1 remaining after PKC activation was determined using quantitative immunoblotting.

Our preliminary results indicate that blocking the lysosomal and proteasomal degradation pathways do not prevent NKCC1 degradation during PKC activation. We have also determined conditions which cause filamentous growth in C.albicans, and will continue searching for others external cues that induce this shift.

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