Manifestation of Novel Social Challenges of the European

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Manifestation of Novel Social Challenges of the European Union in the Teaching Material of

Manifestation of Novel Social Challenges of the European Union in the Teaching Material of Medical Biotechnology Master’s Programmes at the University of Pécs and at the University of Debrecen Identification number: TÁMOP-4. 1. 2 -08/1/A-2009 -0011

Manifestation of Novel Social Challenges of the European Union in the Teaching Material of

Manifestation of Novel Social Challenges of the European Union in the Teaching Material of Medical Biotechnology Master’s Programmes at the University of Pécs and at the University of Debrecen Identification number: TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Tímea Berki and Ferenc Boldizsár Signal transduction B-CELL RECEPTOR SIGNALING

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 B-cell development • Haematopoietic development is a highly regulated

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 B-cell development • Haematopoietic development is a highly regulated multistep process in which pluripotent HSC differentiate through intermediate progenitors to mature cells in the blood • These processes are regulated by transcription factors and signaling pathways • The generationof lymphoid progenitors depends on: c. Kit, Flt 3, and IL-7 R

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Early lymphopoiesis depends on: • PU. 1: crucial for

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Early lymphopoiesis depends on: • PU. 1: crucial for myeloid and lymphoid progenitors • IKAROS: controls development of lymphoid progenitors • Bcl 11 a: zinc finger transcrition factor, in its absence development is blocked in CLP • E 2 A: helix loop helix protein, B-cell fate determinant, turns on Bcell specific genes • EBF: early B-cell factor, B-cell fate determinant, turns on B-cell specific genes • Pax 5: in its absence cells are blocked at pro-B stage, self renew, broad developmental potential. Pax 5 represses non-B genes

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 The stage specific crucial events involved in B lymphoid

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 The stage specific crucial events involved in B lymphoid commitment LMPP Lymphoid priming CLP B-lineage priming BCP B-lineage commitment E 2 A/IKAROS/PU 1 E 2 A/EBF 1/PAX 5 RAG 1/2, Td. T, Ig. H, IL-7 Rα, Notch-1, Ebf 1 CD 19, Tcf 4, Aiolos, Irf 4, Irf 8, CD 55 λ 5, Vpre. B 1, mb 1, B 29, Oca. B, Pax 5 LMPP: lymphoid primed multipotent progenitor, CLP: common lymphoid progenitor, BCP: B lineage committed progenitor

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Genetic control of lineage commitment in early lymphopoiesis Low

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Genetic control of lineage commitment in early lymphopoiesis Low level of PU. 1 → IL 7 Ra expression → lymphoid lineages (IL 7 R +) Pre. DC Pre. NK Id 2 Ikaros HSC PU. 1 low IL-7 R + CLP EA 2 EBF PU. 1 high Notch IL-7 R Myeloid prog. Pro-T Pax 5 Pro-B

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Early B-cell development in the bone marrow Ikaros c-Kit

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Early B-cell development in the bone marrow Ikaros c-Kit Fit 3 PU. 1 (DH ► JH) HSC ELP Myeloid Erythroid Bcl 11 a Il-7 R E 2 A EBF Pax 5 Foxp 1 DH ► JH VH ► DHJH Pre-BCR+ CLP Prepro-B Pro-B Large pre-B T NK VL ► JL Small pre-B BCR+ Imm. B A schematic diagram of early B lymphopoiesis, showing the successive differentiation stages and the rearrangement status of Ig. H and Ig. L genes

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 The Ig gene rearrangement • DH–JH rearrangements are initiated

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 The Ig gene rearrangement • DH–JH rearrangements are initiated in the earliest lymphocyte progenitors (ELPs) at a low level and are completed as the cell progresses to the CLP and pre-pro-B cell (also referred to as CLP 2) stage. • VH–DJH recombination takes place in pro-B cells, and successful rearrangement leads to expression of the Igm protein as part of the pre-BCR in large pre-B cells. • Subsequent rearrangement of the Ig. L locus in small pre-B cells results in the expression of the BCR (consisting of m heavy and k or l light chains) on immature B cells (Imm. B). • The approximate points of the developmental arrest in mice that have defective transcription factors PU. 1, Ikaros, Bcl 11 a, E 2 A, EBF, Pax 5 and Foxp 1 (black) or signalling components involved in signalling through c-Kit, Flt 3 or IL 7 R (grey) are indicated above. • E 2 A- and EBF-deficient progenitors in the bone marrow resemble pre-pro-B cells but lack all Ig rearrangements.

Expression pattern of transcription factors involved in B cell commitment and differentiation Progenitor CMLP,

Expression pattern of transcription factors involved in B cell commitment and differentiation Progenitor CMLP, CLP PU. 1 E 2 A EBF Ikaros Pax 5 LEF-1 NF-k. B Aiolos Bcl-6 Blimp-1 XBP-1 Commitment to B lineage Bone marrow Pre. B → Imm. B Peripheral maturation Germinal center TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Plasma cells

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Immunreceptor Tyrosin-based Activation Motif Activating receptors Immunreceptor Tyrosin-based Ativation

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Immunreceptor Tyrosin-based Activation Motif Activating receptors Immunreceptor Tyrosin-based Ativation Motif (ITAM): ITAM D/E-x 2 - Yxx. L/I-x 6 -9 Yx 2 L/I ITAM ACTIVATION ITAM Inhibiting receptors Immunreceptor Tyrosin-based Inhibition Motif (ITIM): I ITIM L/V/S-Yxx. L/V ITAM INHIBITION ITIM

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Acute antigen signaling Antigen BCR Pl. P 2 Ig

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Acute antigen signaling Antigen BCR Pl. P 2 Ig Pl. P 3 Igβ PI 3 K P P P P SYK LYN BTK PLC 2 BLNK VAV GRB 2 SOS DAG PKC IP 3 Ca 2+ RAS ERK Transcription factors Proliferation, activation and antibody secretion

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Co-stimularory pathways of Bc. R signaling BCR CD 19

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Co-stimularory pathways of Bc. R signaling BCR CD 19 BAFFR Plasma membrane PIP 2 LYN Ig PIP 3 PI 3 K Igβ TRAF 3 ITAM P TRAF 3 AKT 1 P SYK PLC 2 BLNK TRAF 2 DAG IP 3 PKCβ GRB 2 RAS Noncanonical NF- B 1 pathway NIK BTK CARMA 1 Ca 2++ MALTI RAF BCL 10 IKK NFAT Canonical NF- B 1 pathway IKKβ p 50 p 100 RELB p 52 RELB MEK IKK ERK p 65 NF- B 2 NF- B 1 Cytoplasm REL Nucleus P 100 Anti-apoptotic proteins Such as BCL-XL and MCL 1

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Long term Bc. R stimulation BCR CXCR 4 Pl(4,

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Long term Bc. R stimulation BCR CXCR 4 Pl(4, 5)P 2 Ig BAFFR Pl(3, 4, 5)P 3 Igβ P P P PI 3 K Pl (3, 4)P 2 SYK P P LYN SHIP 1 LYN BTK PLC 2 BLNK VAV GRB 2 Migration SOS DAG PKC IP 3 Ca 2+ RAS ERK DOK Transcription factors ? Survival Proliferation, activation and antibody secretion

The positive (CR 2) and negative (Fcg. RIIb) B cell regulation model TÁMOP-4. 1.

The positive (CR 2) and negative (Fcg. RIIb) B cell regulation model TÁMOP-4. 1. 2 -08/1/A-2009 -0011 antigén ant igén C 3 d antigén CD 21(CR 2) CD 19 BCR CD 81 PIP 3 P SH IP Lyn P P P SH IP SH P-2 P a b ab Syk Lyn Syk P Vav Gab 1 PIP 3 Ras/MAPK P P P PI 3 -K

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Role for lipid rafts in B-cell activation CD 45

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Role for lipid rafts in B-cell activation CD 45 CD 22 BCR Ig /β Lipid raft LYN ITIM ITAM LYN Antigen binding CD 45 CD 22 BCR Ig /β LYN Receptor downregulation LYN Internalization P P P P SYK Antigen targeting SYK Signal transduction

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Lipid rafts • The plasma membrane is composed primarily

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Lipid rafts • The plasma membrane is composed primarily of sphingolipids, (glycerol)phospholipids and cholesterol. • Sphingolipids differ from most phospholipids in that they have long, largely saturated acyl chains that allow them to pack tightly in a bilayer, forming a gel phase in which there is very little lateral movement or diffusion. • The gel phase of the sphingolipids is altered by the association of cholesterol, which condenses the packing of the sphingolipids by occupying the spaces between the acyl chains. • So, cholesterol-containing sphingolipid microdomains exist in a liquid-ordered phase that is significantly more fluid than the gel phase.

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Overview of Bc. R signaling BCR Antigen CRAC channel

TÁMOP-4. 1. 2 -08/1/A-2009 -0011 Overview of Bc. R signaling BCR Antigen CRAC channel CD 19 Ca 2+ Fc RIIB 1 Lipid raft aggregation Ig BCR Internalization Gab Igb P 13 K p 110 BCAP Shc PIP 3 GRB 2 p 85 SHIP 2 SHIP 1 Cbl Bam 32 clathrin Lyn CD 22 Syk ezrin SHIP 1 Pl(4, 5)P 2 PIR-B SHIP 2 Nck BLNK PRK 2 Rac/ cdc 42 Rho CD 19 PIP 3 Rac LAB DAG Nucleus PKC Ca 2+ MEK 1/2 JNK 1/2 Erk 1/2 p 38 JNK Erk 1/2 Jun GSK-3 CARMA 1 Rheb Bcl 10 Ca. M m. TOR IKK Ca 2+ ATF-2 ATP generation ER TSC 2 TAK 1 MKK 4/7 p 38 CREB Glycolysis Akt Fcg. RIIB 1 Proteasomal degradation CD 40 Cytoplasm DAG Dok-1 c-RAF PKC Intracellular Ca 2+ store MALT 1 Rap CD 19 MKK 3/4/6 IP 3 R Ras GRP Ras GAP Ras MEKKs IP 3 Bam 32 SOS Riam Glucose uptake Ca 2+ BLNK GRB 2 Rap. L Dok-3 STIM 1 Btk CD 45 PLC 2 Cytoskeletal rearrangments and integrin activation Bam 32 Lyn CD 19 VAV PTEN Rho. A Bcl-6 Egr-1 NF- B Transcription Bcl-x. L Bfl-1 P 70 S 6 K l B Elk-1 Calcineurin NF- B NFAT Ca. MK Oct-2 Ets-1 NFAT Protein synthesis Growth arrest, apoptosis Akt Transcription Fo. XO