Below is a brief summary of some of the current projects in our lab and some of the tools we use to unravel human immune function.
PATients with MONOGENIC IMMUNE Disorders
We recruit patients with rare, inherited immune disorders in order to study the genetic, molecular, and cellular basis of disease. By understanding the mechanisms of immune dysfunction, we aim to devise precision medicine approaches to treat these patients while at the same time gleaning insights into biology that is fundamental for proper immune function in humans. Although we work predominantly with primary human samples, we complement these studies with in vivo mouse models as needed. Moreover, we are developing tissue modeling tools to advance our mechanistic analyses and directly test pharmacological manipulation of immune cell function at tissue sites.
A major focus has been on PASLI/APDS (Activated PI3Kdelta Syndrome), a primary immunodeficiency caused by germline gain-of-function mutations in the PIK3CD or PIK3R1 gene encoding a subunit of the phosphoinositide 3-kinase delta (PI3Kdelta) complex. We have identified multiple distinct mutations sites that all result in recurrent sinopulmonary infections, lymphoproliferative disease, and susceptibility to herpesviruses and B cell lymphomas. Identification of the underlying genetic cause of PASLI/APDS has led to exciting clinical trials of specific PI3Kdelta inhibitors in this disease.
T cell SenesCence
We discovered that PASLI/APDS patients have an abnormally increased frequency of terminally differentiated, senescent T cells, which may contribute to their poor control of herpesviruses. T cell senescence occurs when the DNA damage response (DDR) is activated, which can be triggered by direct DNA insult or extreme shortening of telomeres that are recognized as a double-stranded DNA break. T cells from the blood of PASLI/APDS patients exhibit markedly short telomeres, and our ongoing efforts are directed at better understanding regulation of telomere length by PI3K and related pathways, inflammation associated with senescent T cells, and signal transduction through DDR pathways in T cells.
T cell Death
T cells from APDS/PASLI patients are also more sensitive to T cell receptor (TCR) restimulation-induced cell death (RICD), which can be modulated by acute inhibition of PI3K signaling. Ongoing research is aimed at elucidating the contribution of T cell skewing toward terminal effectors versus hyperactive PI3K signal transduction upon TCR restimulation in this cell death response, which may further contribute to immunodeficiency in APDS/PASLI.
Our lab employs the use of genomics to identify germline mutations in patients with rare and extreme immune phenotypes. Using whole-exome and whole-genome sequencing, we assess affected (usually pediatric) patients as well as their healthy family members to filter for genetic variants in genes that may regulate immune function. Based on our family analyses, we conduct mechanistic laboratory research to uncover cellular and biochemical defects underlying disease.
The immune system is composed of a variety of different cells expressing different surface or intracellular markers. The composition of all these cells and the markers expressed can provide insightful information to how genetic variants may affect these compositions. Flow cytometry and mass cytometry are powerful single-cell technologies used to measure cell populations, changes in surface and intracellular protein levels, and signal transduction responses (e.g., phosphoflow).
Molecular biology and biochemistry
To gain molecular resolution in our analyses of human immune cell function, we use molecular biology to clone, mutate, and overexpress proteins of interest and biochemistry to analyze protein function through approaches including western blotting, co-immunoprecipitation, analysis of post-translational modifications and protein stability, etc..
The Lucas Lab uses confocal and super-resolution microscopy to image both live and fixed cells in order to determine localization and co-localization of proteins, lipids, and organelles.
To extend our studies of signaling in human immune cells, we utilize unbiased proteomics approaches through mass spectrometry. This enables identification of novel protein-protein interactions, kinase substrates, and post-translational modifications in proteins and pathways of interest.