Acute lower respiratory tract infections represent the greatest burden of disease worldwide, with mortality rates that have remained virtually unchanged since the discovery of antibiotics. The effective immune response requires the tightly regulated production of cytokines and chemokines that can facilitate maximal bacterial clearance, while minimizing tissue damage. A better understanding of cytokine regulation in the lung is needed, as future therapeutics targeting the immune response may be an essential given the rising rates of antibiotic resistance. As a foundation for future therapeutics, our laboratory is interested in identifying novel regulators of cytokine elaboration. A major regulator of gene expression, PIWI proteins associate with PIWI- interacting RNAs (piRNAs) in the mammalian germ line, and function to repress ancient retroviral elements. Ablation of any of the three PIWI proteins in mice (MIWI, MILI, or MIWI2) results in defects in spermatogenesis due to aberrant expression of LINE elements. While the germ cell functions of PIWI proteins have been described, next to nothing is known about their potential role(s) in somatic cells. To our surprise microarray analysis and qRT-PCR of sorted alveolar epithelial cells demonstrated that a single PIWI protein, Miwi2 is induced in the lung during bacterial pneumonia. Both in vitro and in vivo depletion of Miwi2 indicates that it acts to promote cytokine and chemokine expression under relevant inflammatory conditions. To our knowledge, this is the first demonstration of a somatic cell function for Miwi2. Given these findings, several important questions still remain. We will test the central hypothesis that during bacterial pneumonia, the piRNA-binding protein Miwi2 is repurposed from its known function as a germ cell specific repressor of ancient viral elements and is induced in pulmonary epithelial cells to promote cytokine expression and immune defense.
In Specific Aim 1 we will use a multicolor FACS strategy to identify the specific alveolar epithelial cell population where Miwi2 is induced during pneumonia.
In Specific Aim 2 we begin to elucidate the molecular mechanisms of Miwi2 dependent cytokine induction.
In Specific Aim 3 we test the hypothesis that Miwi2 dependent expression of cytokines and chemokines are necessary for host defense in a clinically relevant model of bacterial pneumonia. Results of these studies will shed light on the under-described somatic functions of piRNA binding proteins, as well as enhance our understanding of cytokine regulation during bacterial pneumonia. As piRNA binding protein induction has been associated with certain cancers, we anticipate that these data will extend beyond our current focus of innate immunity. Hopefully, the results gained here will serve as a basis for future immunomodulatory therapeutics that improves the outcomes of patients with pneumonia.
The goal of this study is to elucidate novel mechanisms of epithelial cytokine regulation during bacterial pneumonia. We have identified that the normally germ line expressed piRNA binding protein Miwi2 is induced in alveolar epithelial cells during Pneumococcal pneumonia, and serves to promote cytokine and chemokine expression. The proposed studies will provide a deeper understanding of immune gene expression, which is a necessary foundation for the development of future immunomodulatory therapeutics directed at improving outcomes after acute lung injury.
| Wasserman, Gregory A; Szymaniak, Aleksander D; Hinds, Anne C et al. (2017) Expression of Piwi protein MIWI2 defines a distinct population of multiciliated cells. J Clin Invest 127:3866-3876 |
