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Probing the subcellular distribution of phosphatidylinositol reveals a surprising lack at the plasma membrane.

The Journal of cell biology (2020-03-27)
James P Zewe, April M Miller, Sahana Sangappa, Rachel C Wills, Brady D Goulden, Gerald R V Hammond
ABSTRACT

The polyphosphoinositides (PPIn) are central regulatory lipids that direct membrane function in eukaryotic cells. Understanding how their synthesis is regulated is crucial to revealing these lipids' role in health and disease. PPIn are derived from the major structural lipid, phosphatidylinositol (PI). However, although the distribution of most PPIn has been characterized, the subcellular localization of PI available for PPIn synthesis is not known. Here, we used several orthogonal approaches to map the subcellular distribution of PI, including localizing exogenous fluorescent PI, as well as detecting lipid conversion products of endogenous PI after acute chemogenetic activation of PI-specific phospholipase and 4-kinase. We report that PI is broadly distributed throughout intracellular membrane compartments. However, there is a surprising lack of PI in the plasma membrane compared with the PPIn. These experiments implicate regulation of PI supply to the plasma membrane, as opposed to regulation of PPIn-kinases, as crucial to the control of PPIn synthesis and function at the PM.

MATERIALS
Product Number
Brand
Product Description

Supelco
Hydrogen Peroxide Solution 30% W/W, meets analytical specifications of USP/NF GR ACS
Sigma-Aldrich
Albumin, Bovine Serum, Fraction V, Fatty Acid-Free