Author ORCID Identifier
0000-0002-6279-3590
Date of Graduation
8-2017
Document Type
Thesis (MS)
Program Affiliation
Biomedical Sciences
Degree Name
Masters of Science (MS)
Advisor/Committee Chair
Ilya Levental
Committee Member
Yong Zhou
Committee Member
Mikhail Bogdanov
Committee Member
Alemayehu Gorfe
Committee Member
Neal Waxham
Abstract
Mammalian cells maintain a distinct disparity in lipid composition between the two leaflets of the bilayer of the plasma membrane. This compositional asymmetry is most prominent for phosphatidylserine (PS), a negatively charged lipid that is found almost exclusively on the cytoplasmic (inner) leaflet of the plasma membrane. This energetically unfavorable asymmetry is maintained by the activity of ATP-dependent transporters called flippases and destroyed by energy-independent lipid channels called scramblases. Although this compositional asymmetry has been known for decades, there has been little investigation of its structural impact on the physical properties of the membrane, nor its functional impact in healthy cells. Here, we seek to determine the effect of compositional asymmetry on the biophysical properties of the plasma membrane and to determine the function of PS scrambling in the activation of immune cells. Canonically, PS is exposed to the extracellular (outer) leaflet of the plasma membrane in an irreversible manner when cells undergo apoptosis, marking them for clearing by macrophages. The primary mediator of PS exposure in cells (otherwise known as phospholipid scrambling) is a calcium activated chloride channel protein known as TMEM16F (or Anoctamin 6; Ano6). The scrambling activity of this channel is gated by intracellular calcium, with increased Ca2+ levels leading to channel opening and rapid movement of PS from the inner leaflet to the outer. During cell death and platelet activation, this process is irreversible; however, it has been recently shown that it also occurs in immune cells, but reversibly. Although such behavior has been observed across many different immune cell types, so far, no functional role has been attributed to it. Through pharmacological inhibition and genetic deletion of Ano6, we show that plasma membrane asymmetry may affect calcium mediated cell signaling in immune cells. This work paves the way for future studies to determine the broad roles of compositional and biophysical plasma membrane asymmetry in cellular physiology, and the specific mechanisms by which it is involved in various cell signaling pathways.
Keywords
Lipid, asymmetry, Ano6, scramblase, GPMV