p art i immnoglobulins are proteins proteins are specified by genes there are too few genes to...
TRANSCRIPT
IMMUNOLOGY
Bios 328a textbook-based study of immunologySpring 2003
http://www.lehigh.edu/~sk08/Courses/Bios328/mainpage.htm
PART I
• Immnoglobulins are proteins• Proteins are specified by genes• There are too few genes to specify all the
antibodies.– i. e., ~32,000 genes < 10,000,000,000 Ab’s
• How is Ig diversity specified genetically?
Ig proteins are specified by genetic “cassettes”
• Light chains are specified by “variable” (V), “joining (J), and “constant” (C) gene segments (aka “cassettes”).
DNA rearrangementand
alternative RNA spicing
Ig proteins are specified by genetic “cassettes”
• Heavy chains are specified by “variable” (V), “diversity” (D), “joining (J), and “constant” (C) gene segments (aka “cassettes”).
DNA rearrangementand
alternative RNA splicing
Another view….
PART II
• Cassettes rearrange… • How does this happen?
• How do you get one “V” fusing to one “J” (in a light
chain)?• In a heavy chain, a “D” fuses with a “J”; then the fused DJ
cassette fuses with a “V” cassette…
• The orderliness of this process implies that there are genetic instructions. What are they?
Cassettes rearrange…
• The heptamer is a palindrome– (i.e., it exhibits two-fold rotational symmetry.)
• The nonamer is AT-rich
– “Turns” refer to the DNA helix…
Cassettes rearrange…
One turn – two turn rule…
• one turn and two turn are “recombination signal sequences”
• one turn only reacts with a two turn
• Recombination signal sequences are the substrates of enzymes RAG-1 and RAG-2
(“RAG” = recombination-activating gene)
So… cassettes are marked by RSS (i.e., they are substrates for recombination.)
Thus, cassettes can be fused.
What is the consequence?
Look at mouse:
A mouse has:
134 VH, 13 DH, 4 JH segments
85 V, 4 Jsegmentsand 2 V, 3 J segments
Thus, a mouse has:
134 13 4 = 6968 heavy chains 85 4 = 340 kappa chain
and 2 3 = 6 lambda chains
6968 (340 + 6) = 2,410,928 antibodies
PART III (the HARD part…)
2.4 106 < 1010
So, there must be additional mechanisms of diversity other than “fusing” “cassettes”
How does a RAG enzyme work?
Junctional flexibility
The “hairpin loop”
Junctional flexibility, “P” nucleotides, and “N” nucleotides are added to CDR3
Somatic hypermutation
One turn – two turn rule…• one turn only
reacts with a two turn
• crossover between direct repeats (same transcriptional orientation) leads to deletion
• crossover between indirect repeats leads to inversion
PART IV• How do immunoglobulins assemble?
• Some immunoglobulins are in the surface membrane of immature B-cells while other immunoglobulins of the same idiotype are secreted by mature B-cells. What’s the difference?
• Similarly, identical variable regions can be shared among different isotypes. How?
• B-cells are diploids with two sets of genetic instructions. How does just one set get expressed?