We identified Klf15 as a novel target gene of glucocorticoids in airway smooth muscle (ASM) and subsequently found that Klf15–/– mice have an isolated reduction in airway hyperreactivity (AHR) in a model of allergic airway disease. We hypothesize that Klf15 impacts AHR through directly modulating ASM phenotype/function. We are testing this hypothesis by manipulating the expression levels of Klf15 and performing in vivo and in vitro analysis of ASM function/phenotype. Particular areas of focus are: (A) determining the smooth muscle specific effects of Klf15 on AHR in vivo through using smooth muscle restricted Klf15 knockout mice, and smooth muscle specific Klf15 transgenic mice, and (B) establishing mechanisms through which Klf15 over-expression reduces ASM proliferation and blocks ASM hypertrophy in vitro.
We found that glucocorticoids induce a program of gene expression changes that dramatically expands over time both in murine lung and in cultured airway epithelial and smooth muscle cells. We also found that a number of transcription factors are among the early targets of GR signaling. These data led us to define a novel feed forward circuit architecture involving GR and Klf15 that controls the magnitude and timing of expression changes of ~5% of the GR-regulated transcriptome. In addition to completing the molecular characterization of GR:KLf15 feed forward circuits, we are testing the general hypothesis that early GR-regulated transcription factors play key roles in controlling expression changes of subsets of genes in the GR-regulated transcriptome at later time points. We are approaching this research topic through gene knockdown and over-expression, global expression analysis (e.g. RNAseq and micro-arrays), high throughput qPCR, and measuring the generation of nascent RNA.
I previously developed a fluorescent protein-based method to measure four different cellular outputs simultaneously in individual wells of a 96 well dish. My lab now has an ongoing interest in applying this methodology in proof-of-principle high throughput screens to discover compounds that may improve the treatment of lung disease. In one area, in collaboration with Dr. Hong Wei Chu, we are characterizing mucus production in H292 cells, a lung epithelial model, as a basis to (1) develop a screen that probes multiple steps in the mucus production pathway and (2) apply systems biology to analyze mucus regulation. In another screening project, we will develop a system to discover compounds that reduce the risk of glucocorticoid-induced osteoporosis without altering anti-inflammatory effects.
Research in the Gerber Laboratory is supported through institutional funds, a Giles. F. Filley Award, and R01HL109557 (4/15/12 – 3/31/17)