miR-29-TRAF4 axis is a novel regulator of CD40 signaling in malignant B cells

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F65269705%3A_____%2F21%3A00074948" target="_blank" >RIV/65269705:_____/21:00074948 - isvavai.cz</a>

Výsledek na webu

<a href="https://cancerres.aacrjournals.org/content/81/13_Supplement/2368#" target="_blank" >https://cancerres.aacrjournals.org/content/81/13_Supplement/2368#</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1158/1538-7445.AM2021-2368" target="_blank" >10.1158/1538-7445.AM2021-2368</a>

Jazyk výsledku

angličtina

Název v původním jazyce

miR-29-TRAF4 axis is a novel regulator of CD40 signaling in malignant B cells

Popis výsledku v původním jazyce

The synchronous activation of BCR and CD40 signaling via B-T cell interactions is required for proliferation of normal (Luo et al, 2018) and some malignant B cells, especially in chronic lymphocytic leukemia (CLL) and follicular lymphoma (FL). In CLL/FL cells, proliferation occurs mainly in lymph nodes, but not in bone marrow or peripheral blood which lack access to proper B-T cell interactions. Here we have analyzed the mRNA and miRNAs profile in the proliferative intraclonal CLL cell subpopulation that has recently exited lymph node niches (CXCR4dimCD5bright cells; Calissano et al,2011; Pavlasova et al,2016) to reveal molecules potentially participating in synchronous regulation of CD40 and BCR pathway. This has identified 36 miRNAs and 1370 mRNAs differentially expressed in CLL cells exiting lymph nodes as compared to resting non-proliferative CLL cells (CXCR4brightCD5dim cells). Next, we overlapped the 36 miRNAs with their predicted target mRNAs with putative function in CD40/BCR (TargetScan, KEGG) which were anti-correlated to at least one miRNA (miRNAs typically decrease mRNA stability). This revealed among others anti-correlation of lower miR-29 with higher TRAF4 levels in immune niches, which was validated in paired lymph node biopsies vs peripheral blood CLL cells (P&lt;0.01). The negative regulation of TRAF4 by miR-29 was confirmed by transfection of primary CLL and MEC1 cells with synthetic miR-29, miR-29 inhibitor and by a luciferase assay with binding site from TRAF4 3&apos;UTR. Several other miR-29 targets involved in other pathways have been identified as additional targets. TRAF4 is an understudied member of a protein family regulating receptor-induced immune cell activation, however, its role in CD40 pathway or its interactome is unknown. Co-immunoprecipitation and mass spectrometry in B cells pre- and post- stimulation by CD40 ligand revealed TRAF4 interaction with members of the CD40 pathway and a novel function in positive regulation of CD40-induced NFκB activity. In B cells, TRAF4 silencing or its over-expression affected IKKα/β phosphorylation following CD40 ligation or co-culture with activated T cells (all P&lt;0.01). Notably, BCR inhibition lead in vivo to miR-29 upregulation by interfering with its negative regulator MYC induced by BCR. This leads to TRAF4 repression and drastically impairs CD40 signaling (P&lt;0.001), which at least partially explains the observed anti-proliferative effects of BCR inhibitors. Altogether, we have described a novel MYC-miR-29-TRAF4 axis that regulates CD40 signaling in B cells, and acts to synchronize BCR activation with CD40 pathway.

Název v anglickém jazyce

miR-29-TRAF4 axis is a novel regulator of CD40 signaling in malignant B cells

Popis výsledku anglicky

The synchronous activation of BCR and CD40 signaling via B-T cell interactions is required for proliferation of normal (Luo et al, 2018) and some malignant B cells, especially in chronic lymphocytic leukemia (CLL) and follicular lymphoma (FL). In CLL/FL cells, proliferation occurs mainly in lymph nodes, but not in bone marrow or peripheral blood which lack access to proper B-T cell interactions. Here we have analyzed the mRNA and miRNAs profile in the proliferative intraclonal CLL cell subpopulation that has recently exited lymph node niches (CXCR4dimCD5bright cells; Calissano et al,2011; Pavlasova et al,2016) to reveal molecules potentially participating in synchronous regulation of CD40 and BCR pathway. This has identified 36 miRNAs and 1370 mRNAs differentially expressed in CLL cells exiting lymph nodes as compared to resting non-proliferative CLL cells (CXCR4brightCD5dim cells). Next, we overlapped the 36 miRNAs with their predicted target mRNAs with putative function in CD40/BCR (TargetScan, KEGG) which were anti-correlated to at least one miRNA (miRNAs typically decrease mRNA stability). This revealed among others anti-correlation of lower miR-29 with higher TRAF4 levels in immune niches, which was validated in paired lymph node biopsies vs peripheral blood CLL cells (P&lt;0.01). The negative regulation of TRAF4 by miR-29 was confirmed by transfection of primary CLL and MEC1 cells with synthetic miR-29, miR-29 inhibitor and by a luciferase assay with binding site from TRAF4 3&apos;UTR. Several other miR-29 targets involved in other pathways have been identified as additional targets. TRAF4 is an understudied member of a protein family regulating receptor-induced immune cell activation, however, its role in CD40 pathway or its interactome is unknown. Co-immunoprecipitation and mass spectrometry in B cells pre- and post- stimulation by CD40 ligand revealed TRAF4 interaction with members of the CD40 pathway and a novel function in positive regulation of CD40-induced NFκB activity. In B cells, TRAF4 silencing or its over-expression affected IKKα/β phosphorylation following CD40 ligation or co-culture with activated T cells (all P&lt;0.01). Notably, BCR inhibition lead in vivo to miR-29 upregulation by interfering with its negative regulator MYC induced by BCR. This leads to TRAF4 repression and drastically impairs CD40 signaling (P&lt;0.001), which at least partially explains the observed anti-proliferative effects of BCR inhibitors. Altogether, we have described a novel MYC-miR-29-TRAF4 axis that regulates CD40 signaling in B cells, and acts to synchronize BCR activation with CD40 pathway.

Druh

O - Ostatní výsledky

CEP obor

—

OECD FORD obor

30204 - Oncology

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2021

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů