Biology subjects

McNitt, D. H.

Publications and source records attributed to McNitt, D. H..

3 recordsLinked to original sources

bioRxiv2026

SAP loss limits anti-insulin atypical B cell activation and pro-inflammatory CD8 T cells despite preserved Tfh responses to protect against type 1 diabetes

SLAM-associated protein (SAP) is required for T follicular helper (Tfh)-B cell interactions that underlie germinal center formation, but it is unclear if SAP governs islet-reactive CD4+ T cell-B cell interactions and downstream pro-inflammatory CD8+ T cell destruction of islets in type 1 diabetes (T1D). To address this question, we utilized the VH125SD.NOD mouse model, whereby 1-3% of all B cells bind insulin. Germline SAP loss in this model led to reduced T1D incidence and impaired germinal center B cell formation, yet did not alter T follicular helper cell formation or phenotype. SAP loss reduced pro-inflammatory and activated insulin-autoreactive B-T interactions and limited anti-insulin B cell proliferation, activation, and upregulation of co-stimulatory molecules otherwise enhanced in the pancreas. Anti-insulin extrafollicular antibody and memory responses following immunization were preserved in VH125SD.SAP-/-.NOD mice, but activated atypical anti-insulin B cell responses were reduced. Ultimately, SAP loss led to reduced pro-inflammatory CD8+ T cell formation and islet-reactive progenitor exhausted CD8+ T cells in pancreata. These data highlight the essential role of SAP in mediating proinflammatory, anti-insulin B-T interactions to support T1D. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=161 SRC="FIGDIR/small/741363v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@1971e3borg.highwire.dtl.DTLVardef@41d72eorg.highwire.dtl.DTLVardef@963004org.highwire.dtl.DTLVardef@2a76b5_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

BCL6 in T cells promotes type 1 diabetes by redirecting fates of insulin-autoreactive B lymphocytes

Currently approved type 1 diabetes (T1D) immunotherapies broadly target T cells and delay but do not fully prevent diabetes development, highlighting the need for more selective targets. Anti-insulin germinal center B cells are uniquely able to present pathogenic insulin epitopes and drive anti-insulin T cells to adopt a T follicular helper fate. T cell expression of BCL6, a key transcriptional repressor in the germinal center response, is essential for spontaneous diabetes in non-obese diabetic (NOD) mice. However, the impact of T cells on pro-pathogenic anti-insulin B cell activity is still poorly understood. Here, we show that VH125SD.NOD mice with T cell loss of BCL6 still produce peripheral anti-insulin B cells yet are protected against diabetes (relative to Bcl6-sufficient controls). This protection was associated with reduced activation, proliferation, germinal center differentiation, and pancreatic infiltration of insulin-binding B cells. Minimally supervised analysis revealed insulin-binding B cells skew towards atypical memory B cell subsets specifically in pancreas and pancreatic lymph nodes, which was reduced by Bcl6{Delta}CD4 loss. Overall, this work suggests BCL6-expressing T cells are pivotal to license pathogenic insulin-binding B cells. Our findings support BCL6 inhibition as a promising T1D immunotherapy, even after insulin autoimmunity is established in the B cell repertoire. Article Highlights- Loss of floxed Bcl6 via Cd4-Cre protects against type 1 diabetes even when an insulin-skewed B cell repertoire is present - BCL6 loss in T cells reduces anti-insulin B cell upregulation of T cell co-stimulatory molecules, proliferation, and IgG class switching in pancreas and pancreatic lymph nodes in VH125SD.NOD mice - Anti-insulin B cells skew towards atypical and atypical memory B cell phenotypes compared to non-insulin binding B cells in pancreas and pancreatic lymph nodes, only some of which are reduced by T cell loss of Bcl6 - This study highlights the translational potential of targeting BCL6, even after the establishment of insulin-reactive B cells, in line with typical intervention points for at-risk individuals Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/671997v1_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@bf0daforg.highwire.dtl.DTLVardef@11b95fdorg.highwire.dtl.DTLVardef@141ccaorg.highwire.dtl.DTLVardef@6e4100_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2020

Burkholderia collagen-like protein 8, Bucl8, is a unique outer membrane component of a tetrapartite efflux pump in Burkholderia pseudomallei and Burkholderia mallei

Bacterial efflux pumps are an important pathogenicity trait because they extrude a variety of xenobiotics. Our laboratory previously identified in silico Burkholderia collagen-like protein 8 (Bucl8) in the Tier one select agents Burkholderia pseudomallei and Burkholderia mallei. We hypothesize that Bucl8, which contains two predicted tandem outer membrane efflux pump domains, is a component of a putative efflux pump. Unique to Bucl8, as compared to other outer membrane proteins, is the presence of an extended extracellular region containing a collagen-like (CL) domain and a non-collagenous C-terminus (Ct). Molecular modeling and circular dichroism spectroscopy with a recombinant protein, corresponding to this extracellular CL-Ct portion of Bucl8, demonstrated that it adopts a collagen triple helix, whereas functional assays screening for Bucl8 ligands identified binding to fibrinogen. Bioinformatic analysis of the bucl8 gene locus revealed it resembles a classical efflux-pump operon. The bucl8 gene is co-localized with downstream fusCDE genes encoding fusaric acid (FA) resistance, and with an upstream gene, designated as fusR, encoding a LysR-type transcriptional regulator. Using RT-qPCR, we defined the boundaries and transcriptional organization of the fusR-bucl8-fusCDE operon. We found exogenous FA induced bucl8 transcription over 80-fold in B. pseudomallei, while deletion of the entire bucl8 locus decreased the MIC of FA 4-fold in its isogenic mutant. We furthermore showed that the Bucl8 pump expressed in the heterologous Escherichia coli host confers FA resistance. On the contrary, the Bucl8 pump did not confer resistance to a panel of clinically-relevant antimicrobials in Burkholderia and E. coli. We finally demonstrated that deletion of the bucl8-locus drastically affects the growth of the mutant in L-broth. We determined that Bucl8 is a component of a novel tetrapartite efflux pump, which confers FA resistance, fibrinogen binding, and optimal growth. Author SummaryBurkholderia pseudomallei and Burkholderia mallei are highly infectious and multidrug resistant bacteria that are classified by the National Institute of Allergy and Infectious Diseases as Tier one select agents partly due to the intrinsic multidrug resistance associated with expression of the efflux pumps. To date, only few efflux pumps predicted in Burkholderia spp. have been studied in detail. In the current study we introduce Bucl8, an outer membrane component of an unreported putative efflux pump with a unique extended extracellular portion that forms a collagen triple helix and binds fibrinogen. We demonstrate Bucl8s role in fusaric acid resistance by defining its operon via bioinformatic and transcriptional analyses, as well as by employing loss-of-function and gain-of-function genetic approaches. Our studies also implicate the Bucl8-associated pump in metabolic and physiologic homeostasis. Understanding how Bucl8 efflux pump contributes to Burkholderia pathology will foster development of pump inhibitors targeting transport mechanism or identifying potential surface-exposed vaccine targets.

microbiology↗