Why does the acyl chain composition of phosphoinositides matter?

Phosphoinositides are a family of phospholipids that play important roles in regulating the function of proteins that interact with different cell membranes. They are involved in signal transduction, membrane identity and protein trafficking between compartments. Phosphoinositides are known to regulate proteins by the binding of their differentially phosphorylated headgroups to conserved domains in protein effectors (eg PH, FYVE and PX domains), thereby regulating their localization and function. Most phospholipid classes are highly heterogeneous with respect to the composition of their aliphatic chains, but mammalian phosphoinositides are unusually homogenous, most reports suggesting they are >75% stearoyl/arachidonoyl (‘C38:4’; total carbons:double bonds). However, the evolutionary driving force for this selection is completely unknown.
The enzyme PI3K phosphorylates one phosphoinositide called PIP2, into another called PIP3, in the inner leaflet of the plasma membrane. PI3K is activated by cell surface receptors for most growth factors and the PIP3 that is produced activates a signalling network that co-ordinates cell growth and movement. The importance of this pathway is underscored by the number of highly prevalent mutations in this pathway which occur in many human cancers and several human overgrowth syndromes, including the frequent deletion or inactivation of a phosphoinositide phosphatase called PTEN.
During our recent work attempting to understand the role of PTEN in a mouse model of prostate cancer (see Malek et al), we used our mass spectrometry methods (see Clark et al) to measure large increases in the levels of phosphoinositides in PTEN-null prostate tissue. To our surprise, we noticed that the increased PIP3 in PTEN-null prostate was a different molecular species to that normally found in differentiated mammalian tissues (unpublished).
This project is aimed at understanding how the loss of PTEN causes a different molecular species of PIP3 to accumulate and the differential properties this new molecular species may bring to the PI3K signaling network.
Plan
Establish the molecular composition of different phosphoinositides in several different mouse tissues, including normal and PTEN-null mouse prostate. This will include MALDI mass spec imaging of tissue slices (in collaboration with AZ), to give cell-type-specific resolution in vivo, hugely extending what has currently been described.
Establish prostate cell lines or organoids which replicate PTEN-dependent acyl chain remodelling. If this fails, we may need to go to model cell lines.
Use isotope tracers in conjunction with CRISPR/Cas9 editing to define the PTEN-sensitive points in phosphoinositide metabolism that lead to acyl chain remodelling.
Attempt to engineer cells in which the molecular composition of phosphoinositides is altered and then study the effects on phosphoinositide function, including PLC and PI3K signaling pathways.

Training
Cutting edge lipid mass spectrometry.
Cell and molecular biology, including cell culture, fluorescent imaging, CRISPR/Cas9 editing
Mouse models of cancer
Intracellular signaling pathways that regulate cell hypertrophy and transformation
Close collaboration between academic (PTH/LS) and pharmaceutical groups (SC).

Impact
Understanding the significance of acyl chain composition to phospholipid function.
Potential to discover novel vulnerabilities in cancer cells.