Elucidating roles for nono in humoral immune… (F32AI200158) – Project Grant to Northwestern University

Abstract: Non-POU-domain-containing octamer binding protein (nono) is a member of a family of proteins (drosophila behavior/human splicing, dbhs) capable of interacting with DNA, RNA and protein. Dbhs proteins function as molecular scaffolds, bridging these interactions to facilitate transcriptional and post-transcriptional regulation in addition to other essential biological processes. A large body of literature has focused on the role of nono in the formation of mammalian-specific, nuclear membraneless organelles known as paraspeckles. These studies have implicated paraspeckles as transcriptional regulators based on their increased abundance in transformed cancer cell lines and tendency to contain RNA-binding proteins. However, many of the associations that have been made regarding paraspeckles are correlative in nature and so IT is unclear how paraspeckles function. Due to associations with several biological functions, separating nono function from paraspeckle function is essential. The objective of this proposal is to use b cells as a model to investigate paraspeckle function within the broader context of nono. Our preliminary data indicates that nono is highly expressed in b cells and upon stimulation in vitro, b cells form paraspeckles in ~40% of cells. Moreover, stimulation of activated b cells via the b cell receptor leads to increased paraspeckle size and number. Thus, we hypothesize that nono-and in part, paraspeckles-play a role in b cell activation responses and transcriptional regulation during the b cell germinal center response. Previous b cell studies of nono have relied on the use of a germline deletion of nono. Given pleiotropic effects observed in germline deletion of nono, we generated a novel b-cell-specific conditional deletion model for nono (nono KO) in mice. Experiments proposed in aim 1 will use a combination of RNA-SEQ, atac-SEQ and cut & tag analysis of control and nono KO activated germinal center b cells to identify direct targets of nono in b cells. We observed that nono-deficient b cells stimulated in vitro expressed more IGE, which is associated with driving allergy responses in mammals. In aim 2, we will utilize our in vitro culture system in conjunction with an in vivo immunization model to dissect functions of paraspeckle- dependent and -independent function of nono in the generation of IGE b cells. Collectively, these experiments will allow US to rigorously interrogate direct targets of nono activity and attempt to identify a paraspeckle- specific function of nono in b cells. This model will serve as a foundation for future studies interrogating paraspeckle function in b cells. As i transition into an independent investigator, i intend to expand my investigation of paraspeckles into other immune cell subsets, likely t cells, to attempt to connect b cell paraspeckle functions to a broader cellular context.

Elucidating roles for nono in humoral immune responses

Award facts

Description

Quick answers

Who received award F32AI200158?

How much is award F32AI200158 worth?

See this award's full record