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Subramanian Lab Overview

To understand the molecular basis of immune disorders, it is critical to understand the earliest – inborn or innate – events that initiate them and how they cause dysregulation of normal immune networks. That’s what drives my interest in applying a systems approach to understanding innate immunity.

–Assistant Professor, Dr. Naeha Subramanian
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Research Overview:

The innate immune system detects microbial infection by employing a network of pattern recognition receptors that are present on the surface of the cell or in the cytoplasm. Nucleotide-binding domain leucine-rich repeat containing (NLR) proteins form the largest known family of intracellular innate immune sensors whose structural hallmark is a modular organization of domains with distinct function: an N-terminal effector or protein interaction domain that mediates downstream signaling cascades, a central nucleotide oligomerization domain (NOD) and a C-terminal leucine rich repeat (LRR) domain for ligand binding. Their most well-recognized function is as cytosolic sensors for conserved pathogen-associated molecular patterns (PAMPS) and danger-associated molecular patterns (DAMPS). PAMPs recognized by NLRs include muramyl dipeptide (sensed by NOD2), diaminopimelic acid (sensed by NOD1), flagellin (sensed by NLRC4), and viral RNA (sensed by NLRP3). Among the DAMPs sensing NLRs, NLRP3 in particular has been shown to be activated by crystals of monosodium urate and calcium pyrophosphate dehydrate leading to gout, and cholesterol crystals leading to atherosclerosis. Upon activation, some members of the NLR family such as NLRP3, NLRP1, and NLRC4 form a macromolecular signaling complex called the inflammasome that acts as a scaffold for activation of caspase-1 leading to processing and secretion of the potent inflammatory cytokine IL-1β. Others such as NOD1 and NOD2 form signaling platforms that activate NF-κB. Thus the NLR family of cytosolic sensors has recently emerged as an important focus of inflammation research.

Several newly emerging aspects of the biology of NLR proteins point to functions that extend beyond inflammation to include tissue homoeostasis, autophagy, nuclear functions as transcription factors, embryonic development, and reproduction. In addition, gain of function mutations as well as polymorphisms in NLRs are associated with a host of severe human autoinflammatory and autoimmune disorders ranging from rare hereditary periodic fever syndromes (NLRP3), early-onset sarcoidosis (NOD2) to more common disorders such as rheumatoid arthritis (CIITA), vitiligo (NLRP1) and inflammatory bowel diseases (IBD) like Crohn’s disease and ulcerative colitis (NOD1 and NOD2). Some NLRs can negatively regulate a variety of intracellular signaling pathways, but some of these “inhibitory NLRs” can also, under certain conditions, activate inflammatory pathways. However, in the setting of a host response to an insult, the combined outcome of these signaling pathways would collectively determine the quality, magnitude and duration of the ensuing host response. Dr. Naeha Subramanian’s work seeks to apply a systems approach to gain a comprehensive understanding of NLR response phenotypes and associated signaling pathways. The goal is to provide insights into not only how NLRs normally function but to ultimately harness this information for therapeutic intervention in patients suffering from conditions related to aberrant NLR function.

Research Focus:

Naeha’s research seeks to extend investigations into NLR function using a combination of imaging, microarray, proteomic and bioinformatics approaches. In the development phase of her new laboratory research is centered on the following projects:

1. Role of organelles in regulation of NLR function:

Naeha’s previous work has shown that NLRP3 associates with mitochondria upon activation and the mitochondrial adapter protein MAVS plays a stimulus-specific and amplifying role in NLRP3 activation (Subramanian et al, Cell, 153, 348–361, 2013). The NLRP3 inflammasome has been implicated in “sterile inflammation” during chronic inflammatory illnesses with an underlying component of mitochondrial dysfunction, such as Type 2 diabetes, obesity, metabolic syndrome, gout, atherosclerosis and Alzheimer’s disease. Therefore, a deeper understanding of the interplay between NLRP3 and mitochondria is clearly of great value.

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Role of mitochondria in promoting NLRP3 inflammasome activation

Future studies will employ a combination of high-throughput experimental, sub-cellular imaging, proteomic and bioinformatic approaches to further dissect mitochondrial regulation of NLRP3 function and extend these investigations to understanding the role of intracellular positioning to organelles in the regulation of other cytosolic innate signaling pathways.

2. A systems approach to understanding NLR function

In humans, 23 NLRs have been identified. These share similarities in structural organization but have been hypothesized to have distinct biological functions based on sequence and structure modeling analysis. To date, however, only a few NLRs have been studied intensively, and the activating stimuli, physiologic functions, and relevant signaling pathways of most members of the NLR family are unknown or poorly defined. Dr. Subramanian’s long-term goal is to undertake a systems approach to gain a comprehensive understanding of NLR response phenotypes and associated signaling pathways using microarray/RNA-Seq/proteomic analyses and pathway reconstruction to identify the transcriptional profiles, proteome changes and response pathways that are triggered as a result of NLR activation. For this purpose, a system for stable inducible expression of NLRs in the appropriate cell type has been developed. A key aspect will include combining information from multiple levels such as genes, proteins, whole cells, and external activators of NLRs to gain a clear understanding of responses triggered by NLRs, their roles in immunity and to eventually understand how NLRs might behave when perturbed by infectious agents or danger signals.

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A systems approach to deciphering NLR function

 

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