Expression and function of NIK‐ and IKK2‐binding protein (NIBP) in mouse enteric nervous system

Abstract

Background NIBP/TRAPPC9 is expressed in brain neurons, and human NIBP mutations are associated with neurodevelopmental disorders. The cellular distribution and function of NIBP in the enteric nervous system (ENS) remain unknown. Methods Western blot and RT-PCR analysis were used respectively to identify the protein and mRNA expression of NIBP and other neuronal markers. Multilabeled immunofluorescent microscopy and confocal image analysis were used to examine the cellular distribution of NIBP-like immunoreactivity (IR) in whole mount intestine. Enteric neuronal cell line (ENC) was infected with lentivirus carrying NIBP or its shRNA expression vectors and treated with vehicle or TNFα. Key Results NIBP is expressed at both mRNA and protein levels in different regions and layers of the mouse intestine. NIBP-like-IR was co-localized with various neuronal markers, but not with glial, smooth muscular, or ICC markers. A small population of NIBP-expressing cells and fibers in extra-ganglionic and intra-ganglionic area were negative for pan-neuronal markers HuD or Peripherin. Relatively high NIBP-like-IR was found in 35-44% of myenteric neurons and 9-10% of submucosal neurons. Approximately 98%, 87% and 43% of these relatively high NIBP-expressing neurons were positive for ChAT, nNOS and Calretinin, respectively. NIBP shRNA knockdown in ENC inhibited TNFα-induced NFkB activation and neuronal differentiation, whereas NIBP overexpression promoted it. Conclusions & Inferences NIBP is extensively expressed in the ENS with relatively high level in a subpopulation of enteric neurons. Various NIBP expression levels in different neurons may represent dynamic trafficking or posttranslational modification of NIBP in some functionally-active neurons and ultimately regulate ENS plasticity.

Elevated NIBP/TRAPPC9 mediates tumorigenesis of cancer cells through NFκB signaling

“…Four short hairpin RNAs (shRNAs) encoded by 4 different regions targeting the 5′-(NR), 3′-coding region (CR) and 3′-untranslated regions (UTR) of human NIBP (NM_031466) were designed for the cloning into lentiviral shRNA expression vector pLL3.7 and their efficacies were evaluated as we described previously [13, 32]. Using the most effective shRNAs, NIBP-NR and -CR [13], we established breast (MDA-MB-231) and colon (HCT116) cancer cell lines with NIBP stable knockdown.…”

“…Three recent studies added to our finding that NIBP increased TNFα-induced NFκB activation in HEK293T cells, fibroblasts, and neurons [32, 34, 35]. Since NIBP was highly expressed in cancer cells and NFκB/IKK2 signaling is critical for tumorigenesis, we thus investigated whether NIBP affects NFκB signaling in cancer cells.…”

“…In HEK293T cells, PC12 cells, enteric neuronal cell lines and brain tissues, NIBP has been shown to interact with IKK2 but not IKK1 [13, 32]. Such interaction contributes to the enhancing effect of NIBP on cytokine-induced activation of the NFκB canonical pathway [13].…”

“…A decade of studies has identified the important functions of NIBP in regulating NFκB signaling [13-15, 32], trans -Golgi networking [19-22] and virus replication [35]. In the present study, we demonstrated that NIBP is highly expressed in numerous cancer cell lines and tumor tissues, and provided evidence for the first time that NIBP plays an important role in regulating viability, proliferation, migration and tumorigenesis of breast or colon cancer cells, notably in response to extracellular pro-inflammatory stimuli.…”

A homozygous founder mutation inTRAPPC6Bassociates with a neurodevelopmental disorder characterised by microcephaly, epilepsy and autistic features

“…Besides its role in membrane trafficking as part of complex TRAPP II, TrappC9 is also involved in the nuclear factor-kappa B signalling pathway 1 14. This protein has been implicated in neural differentiation,15 and inactivation in PC12 cells prevents nerve growth factor-induced neurite outgrowth 14. Its expression is highest in the human cerebellum, suggesting that TrappC9 may have a brain-specific signalling function outside of its general role in membrane trafficking 16.…”