BIOT 307 MOLECULAR IMMUNOLOGY INNATE IMMUNITY

External – Skin – Epidermis and dermis are protective layers – Other protectants pH ~ 3-5 b/c of sebum  destroys bacteria saturated fatty acids  control fungi Β-defensins, psoriasian, dermicidin Low moisture – Broken when goes into blood vessels in innermost layer or hypodermis, subcutaneous layer

External – Mucous, many pathogens enter here – Mucous is glue-like and traps – Wash away – Contain antibacterial and antiviral substances – Passages end up in stomach – low pH – Pathogen invasion aided by: Surface projections and adhesins

INNATE IMMUNITY Mucosal Tissues – Respiratory – Alimentary – Urogenital Chemical secretions – Lysozyme: digests cell wall of G+ and peptidoglycan on G- – Muraminidase – Gastric juice – Defensins from epithelial cells – Cathelicidins bacteria, fungi, env. virus

Normal microbiota – Protective, block invasion, secrete bacteriocins – Gut, mouth, upper respiratory, urogenital, gastrointestinal tracts – Huge sequencing effort underway to identify these organisms in normal and diseased states, esp. chronic diseases

PATTERN RECOGNITION RECEPTORS (PRRs): One Key to IRR Germ-line encoded receptors = pattern recognition receptors – Target conserved molecules, PAMPS (pathogen associated molecular pattern), on bacteria, viruses, fungi, protozoa not on host LPS, peptidoglycan, dsRNA, zymosan, … – Found Extracellular (secreted) Intracellular compartments/cytosol – all cells of same lineage have same ones

EXTRACELLULAR/SECRETED Mechanism – Bind to cell walls targeting them for recognition by complement and phagocytes AND/OR – Opsonins that enhance phagocytosis Examples: – Mannose binding lectin/protein (MBP) – C-reactive protein (CRP) – Serum amyloid protein (SAP) – LPS-binding protein (LBP) Acute phase PRR proteins made by liver early in infection

Involved in Clotting Two Secreted PRRs: CRP, MBP SAP made in acute phase liver response

NEW CONCEPT DIGRESSION: COMPLEMENT SYSTEM Important mechanism to control infection Promotes phagocytosis Involved in bactericidal mechanisms Mediates inflammation through mast cell degranulation and chemotaxis Promotes lymphocyte function

COMPLEMENT PATHWAYS FIGHT INFECTION Opsonize pathogens – cover with complement proteins  enhances phagocytosis induce inflammatory responses enhance antibody responses attack some pathogens directly

Three Complement Pathways MAC

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BACK TO EXTRACELLULAR/SECRETED PRRs Mannose binding lectin/protein (MBP) C-reactive protein (CRP) Serum amyloid protein (SAP) LPS-binding protein (LBP)

EXTRACELLULAR/SECRETED Mannose binding protein (MBP) – Part of C-type lectin superfamily – Associates with and activates serine proteases: MASP-1 and MASP-2 – After binding to pathogen surface this complex activates lectin pathway of complement system, C2* and C4* *Complement protein naming C2 = complement protein 2, etc.

MB-LECTIN MASP = MBL- associated serine protease MBL mannose

MB-LECTIN

ANOTHER VERSION POINTS OUT TOTALITY OF CLEAVED C3 FUNCTIONS

WHEN ACTIVATED MASP-2 cleaves C4 and C2 MASP-1 cleaves C3 and C2

EXTRACELLULAR/SECRETED C-reactive protein (CRP) and serum amyloid protein (SAP) – Bind to phorphorylcholine (PC) on bacteria, other microorganisms, damaged host cell membranes PC found in teichoic acids, capsular carbohydrates, and lipopolysaccharides Requires Ca ++ – Function directly as opsonins (enhancer of phagocytosis) – Function indirectly by binding to C1q of classical complement pathway and activate complement cascade

w/o Ab

CLASSICAL w/o Ab

NOTE: for all pathways regulation occurs: pathogens do not have ability to inactivate bound C3b

SEE NEXT SLIDE FOR WORD DESCRIPTION

Schematic representation of the classical complement pathway. A.The pathway is initiated by binding of two antibody molecules to a multivalent antigen, followed by binding of complement protein C1q, and then by binding of two molecules each of C1r and C1s to C1q to form the active complex C1 B. C4 binds to the C1q portion of C1 C. esterolytic site on C1s activates C4 by splitting off C4b, which binds the antigen-antibody complex or the nearby cell surface D.The same site on C1s then cleaves and activates C2 which has bound to C4b, yielding a C4b2a complex (C3 convertase) E.The C3 convertase binds C3 and cleaves it to the active form C3b F. which binds to the cell surface and to the C3 convertase to form a C4b2a3b complex (C5 convertase) G. late phase of complement activation begins when the C5 convertase cleaves C5 to the active form C5b. H. C5b remains at the cell surface and binds first C6 then C7; I.C7 is hydrophobic and inserts into the plasma membrane J.where it is joined by C8 K. As many as 19 molecules of C9 then polymerize at C5678 to form pores in the plasma membrane L.this final complex is called the membrane attack complex (MAC). Entry of water through the pores into the cell causes osmotic swelling and cell rupture

EXTRACELLULAR/SECRETED LPS-binding protein (LBP) – Lipid transfer molecule binds to monomeric LPS and to high-affinity LPS receptor named CD14 and, on macrophage, neutrophils, DCs – LBP + bactericidal permeability increasing protein (BPI) binds LPS on bacteria and then to CD14, a high affinity LPS receptor. See later TLR4

Extracellular factor (LPS) carried by LBP to CD14 where it binds to TLR4 and then MD2 binds

Simplified Version

INTRACELLULAR/CYTOSOL Scavenger receptors Mannose receptor Toll-like receptors (TLRs) Nucleotide-binding oligomerization domain (NOD 1, 2) Retinoic acid inducible gene-1 (RIG-1)

INTRACELLULAR/CYTOSOL Scavenger receptors – Glycoprotein PRRs recognize LPS and lipoteichoic acid Intact G- and G+ bacteria Damaged host cells and tissues Apoptotic and senescent cells modified low-density lipoproteins – Six classes

INTRACELLULAR/CYTOSOL Mannose receptor, CD206 – C-type lectin CHO binding protein on macrophages MRC1 and MRC2 – Recognizes and binds to mannose, fucose, N- acetylglucosamine, etc.  engulfment, endocytosis – Enhance Ag acquisition for presentation to TC

Cysteine-rich Fibronectin type 11 C-type CHO recognition domains

To be continued