miRNA-155 function in CD8+ T cells

Question 1

Usherwood et al.

Answer 1

miRNA-155 is critical for generation of memory CD8 T cells during virus infection in vivo

• stimulated OT-1 cells with Ag in vitro –> miRNA-155 peaked at day 3 and declined afterwards (decline more marked in cells cultured with IL-15 vs. IL-2) –> culture with IL-2 generates more effector T cells –> sustained miRNA-155 expression required for effector T cell phenotype and not memory
• Mixed bone marrow chimera mice –> miRNA-155 absent from the entire CD8 T cell compartment –> viral titers much higher in mice lacking miRNA-155 in T cells

Question 2

Gracias et al. The microRNA miR-155 controls CD8+ T cell responses by regulating interferon signaling

Answer 2

Higher miRNA-155 expression in CD8 T cells activated in vitro (24hr with anti-CD3 + anti-CD28 42X, increase on days 3 and 5; TNF, IFNg, IFNb, IL1b did not affect expression) and in antigen-specific effector cell in vivo (Thy-1.1+ OT-1 cells transferred into Thy 1.2+ mice + infected with recombinant influenza strain expressing OVA –> D10 miRNA expression high in CD44+CD62L- cells, still 4X at day 60)

MiRNA-155 was intrinsically required for optimal primary CD8+ T cell response against infection and influenza; important for the proliferation of CD8+ T cells and affected many of its targets assoc. with type I IFN

CD8+ T cell responses require miRNA-155: survival and proliferation

Over-expression of miRNA-155 augments CD8+ T cell response: transduced Thy1.1+OT-1CD8+ T cells with control retroviral MigR1 vector expression GFP or same vector expressing mIRNA-155 + GFP –> transferred into Thy-1.2+ mice

Early TCR signaling? –> calcium flux: loaded cells with fluorescent Ca indicator Fluo4 (NO)

MiRNA-`155 deficiency conferred a proliferative defect on CD8+ T cells and enhanced their susceptibility to to type I IFN mediated inhibition (STAT1 pathway) - synergistic effect of genes

In vitro activated miR-155 deficient CD8+ T cells had higher total STAT1 expression

Question 3

What is SOCS-1?

Answer 3

• mouse gene encodes 212 aa protein (human 211) - 90-95% aa identity; amino-terminal domain+kinase inhibitory region (KIR)+central SH2 domain+carboxy-terminal SOCS box morif
• Mechanism of action: classical neg feedback loop –> cytokine signaling induces SOCS1 protein that inhibits JAK by binding via SH2 domain; directly to phosphotyrosine residues; targeting proteins to the elongin BC complex for ubiquitination
• T cells: inhibitory mechanism that that is induced upon T cell activation –> inhibition of TCR signaling that includes Jak/STAT signaling

Question 4

O’Connell et al. miRNA-155 promotes autoimmune inflammation by enhancing inflammatory T cell development

Answer 4

Mir155 KO mice are resistant to EAE induced by MOG35-55: reduced inflammatory conditions in KO mice

Lethaly irradiate WT mice reconstituted with either MiR-155-/- or MiR-155+/+ BM cells –> 4mo –> engrafment and localization of MiR-155 deficiency –> miRNA155 FUNCTIONS IN THE HEMATOPOIETIC COMPARTMENT TO PROPAGATE EAE

MiR-155-/- mice exhibit defective inflammatory T cell development during EAE –> deficiency in Th1 and Th17 cells

T cell intrinsic role of miR-155 in the development of inflammatory T cells during EAE: BIC and miR-155 expression in CD4+ T cells grown in conditions to promote Th17 cell development