JOURNAL CLUB, 28th september 2007

T-CELL ACTIVATION T-CELLS ARE ABLE TO DISTINGUISH BETWEEN SELF AND NON SELF ANTIGENS (Ag) T-CELL RECEPTOR (TCR) PLAY AN IMPORTANT ROLE, BECAUSE IT RECOGNIZES Ag PRESENTED BY APC IN MHC AND THIS LEADS TO THE INDUCTION OF INTRACELLULAR SIGNALING AND T-CELL RESPONSES T-CELLS RESPONSIVENESS AND TCR SIGNALING VARY WITH T-CELL DEVELOPMENT miRNA WERE RECENTLY DISCOVERED TO BE DIFFERENTIALLY EXPRESSD DURING HEMATOPOIETIC DIFFRENTIATION AND THEY MAY BE INVOLVED IN CONTROLLING THE DEVELOPMENT OF IMMUNE SYSTEM

RESULTS DN (CD4 -, CD8 - ) SP (CD4 -, CD8 + ) (CD4 +, CD8 - ) DP (CD4 +, CD8 + ) IMMATURE T-CELLS DN1 (CD44 +, CD25 - ) DN2 (CD44 +, CD25 + ) DN3 (CD44 -, CD25 + ) DN4 (CD44 -, CD25 - ) ORDER OF APPEREANCE DURING DEVELOPMENT DINAMIC REGULATION OF miR181a EXPRESSION IN T-CELLS MATURATION

miR181a EXPRESSION AUGMENTS TCR SIGNALING TO ACCESS HOW miR181a AFFECTS TCR SIGNALING: INCREASED EXPRESSION OF miR181a IN 5C.C7 Ag PRIMED CD4 + T-CELLS STIMULATED WITH MCC (AGONIST); TCR SIGNALING STRENGHT ASSESSED BY MESASURING Ca2 + INCREASE HIGH LEVEL OF miR181a AUGMENTS TCR-MEDIATED T-CELLS ACTIVATION

miR181a EXPRESSION AUGMENTS TCR SIGNALING TCR SIGNAL INPUT=NUMB OF ANTIGENIC pMHC COMPL. AT T:APC INTERFACE TCR SIGNAL OUTPUT=CALCIUM CONCENTR CHANGES IN CYTOSOL miR181a EXPRESSION INCREASES T CELLS SENSITIVITY

miR181a CONVERTS ANTAGONIST INTO AGONIST MCC99R INDUCE CALCIUM INCREASE IN miR181a CELLS MCC99R ALONE STIMULATE EFFECTOR T-CELLS FUNCTION

miR181a EXPRESSION DOESN’T ALTER CD4 AND TCR DENSITY ON T-CELLS SURFACE WHEN CD28 IS BLOCKED AND miR181a T-CELLS ARE TRETAED WITH MC99R, THEY CAN STILL RESPOND TO THE ANTAGONIST. miR181a MODULATE TCR SENSITIVITY BY CONTROLLING INTRACELLULAR SIGNALING MOLECULES, SUCH AS SHP1 (Lck NEGATIVE REGULATOR) AND ERK1/2 PATHWAYS USING UNPUBLISHED COMPUTER PROGRAMS, THEY FOUND TARGETS FOR mir181a: Lck-SPECIFIC-TYROSIN POHOSPHATASE, SHP-2 AND PTPN22 ERK-SPECIFIC-PHOSPHATASE, DUSP5 AND DUSP6

miR181a REPRESSES MULTIPLE PHOSPHATASE IN T-CELLS

miRNA181a INCREASES THE BASAL PHOSPHORILATION LEVELS OF Lck AND ERK miR181a EXPRESSION INHIBITS Lck/SHP1 INTERACTION

MECHANISM OF TCR SIGNALING

MULTITARGET REPRESSION IS REQUIRED FOR miR181a FUNCTION IN TCR SIGNALING STRENGHT OF Ca 2+ RESPONSE AND % OF T-CELLS ACTIVATED EFFICACY AND SPECIFICITY OF shRNA CONSTRUCTS

CONTRIBUTION OF EACH PHOSPHATASE TO Ag DISCRIMINATION

miR181a MODULATES NEGATIVE AND POSITIVE SELECTION OF T-CELLS TREATMENT WITH ANTAGOMIR RESTORED PHOSPHATASE/KINASE INVOLVED IN TCR SIGNALING IN DP CELLS INHIBITION OF ERK ACTIVITY

CONCLUSIONS miRNAs ARE A NEW CLASS OF REGULATORY MOLECULES IN THE IMMUNE SYSTEM miR181a IS AN INTRINSIC MODULATOR OF TCR SENSITIVITY IN T-CELL DEVELOPMENT (ALTHOUGH OTHER COMPONENTS MAY CONTRIBUTE TO THESE CHANGES IN SENSITIVITY) miRNAs CAN MODULATE MULTIPLE TARGETS AS THEY EXERT A QUANTITATIVE REGULATION THAT ALLOWS A DISCRIMINATIVE SWITHC IN Ag RECOGNITION

Paper discussion  What is the key experiment? (the one confirming the statement in the title)  What is the strongest point?  What is the weakest point? and What to do to strenghten it?  What is the take home message? (summarize it in a sentence)

POSITIVE Any cell that doesn’t recognize self MHC is deleted Ensures functional matching of receptor, co-receptor and class of MHC Lineage commitment NEGATIVE Any cell that reacts strongly to self antigen is deleted