Genetics of Ig and TCR Diversity-Hudig Flashcards
What is infliximab?
it is a humanized monoclonal antibody to TNF alpha that reduces immune responses and inflammation (used for rheumatoid arthritis and chrons)
Do we have enough DNA to encode all the different antiodies and TCR receptors? THen how do we have them?
no, genetics with cassettes for variable regions for both immunoglobulin and T cell receptors. Genetics of the immunoglobulin class switch. somatic mutation of ig antigen binding sites
What is a class switch?
it is when a B cell switches from making IgM to a different kind of Ig but still has the same antigen binding specificity
The receptors of T and B cells have variable regions and the remainder of the receptors are constant regions.
T or F
T
Genes for Ig and TCR contain (blank) for variable regions.
cassettes
Each gene locus for Ig has how many cassettes of minigenes?
2 or 3
What is a cassette?
minigenes for making the light and heavy chains with alternative splicing to get different introns
T or F
both light and heavy chains have constant domains. The constant regions are the same for ALL antibodies of the same isotype
T
How do you make a heavy chain out of the cassettes?
you need a V, a D, and a J
How do you make a light chain?
One V and one J, NO D’s
How many chromosomes have V,D,J cassettes and what are they?
7 For B cells o B cell heavy chain – Vh o B cell lambda light chain – Vlambda o B cell kappa light chain – Vkappa
For T cells o TCR alpha chain - Valpha o TCR beta chain – Vbeta ((((((superantigens bind to V beta))))))))) o TCR gamma chain - Vgama o TCR delta chain – Vdelta
Where do you find the D minigene cassette?
ony Ig H, TCR beta and TCR delta loci
Where can you get somatic mutation in Ig?
in Ig L and H variable regions
Do you get somatic changes in TCR?
NOOOOOO!!!!
only in Ig
What do you need to make the T and B cell receptors and what will you get if you dont have them?
RAG1 and RAG2
(they splice minigene cassettes to make diffferent B and T receptors) without either of these you will get SCID
What do the 7 loci to make the B and C receptors have in common?
They all have variable regions
RAG1 and RAG2 are expressed only in (blank)
developing lymphocytes (thymus and bone marrow)
Tumors that express RAG1 and RAG2 are derived from eary developmental stages of (blank)
lymphocytes
(blank) adds N-nucleotides to the V,D, and J exons during antibody gene recombination, enabling the phenomenon of junctional diversity.
TdT
Tumors that express TdT are also derived from early developmental stages of (blank)
lymphocytes
(blank) is DNA inherited in the embryo and kept unchanged in the gonadal germline cells.
Germline DNA
(blank) is DNA found in differentiated cells. It’s the same as germline DNA for most cells.
Somatic DNA
T and F
For both of the chains of TCRs and immunoglobulin, DNA segments are selected from the germline DNA and the DNA is rearranged (with DNA removed) to form somatic genes that will encode the light and heavy chain proteins.
T
T and B cell receptors are considered somatic DNA why?
because the DNA is messed with, rearranged, spliced, deleted therefore making it different from the germline DNA
How do you get germline rearrangments for B cells?
Casettes of minigenes
some are used and some are deleted. The DNA then becomes rearranged without mutation. This makes for production of different antibodies, different, and different receptors
(blank) and (blank) are necessary fro DNA breaking and rejoinging insides the mini cassettes
Rag 1 and Rag 2 (recombinase-activating gene)
What does TdT do?
allows for the rejoining of mnicassettes after there has been alterations
How do you make a heavy Ig chain?
1 VH minigene, 1 DH minigene, I JH minigene plus CH filled in by TdT to create a heavy Ig chain.
(blank) fills in gaps where DNA was cut and joined. The original DNA retains its original sequence, so TdT fills in new sites so TdT does not cause mutations of the original DNA.
Terminal deoxynucleotide transferase (TdT)
How do you make an antibody?
with one light chain and one heavy chain
Hypermutation at hot spots occurs (before/after) the V region minigenes are rearranged, in the CDR (complementary determining regions)
after
(blank) occurs each time an antigen is re-encountered
somatic mutation
One Ig Heavy chain is made of 1 Vh V minigene, 1 D minigene, 1 J minigene, and a Ch. The (blank) region determines the isotype
The Ch (constant region)
o T and B cells don’t have somatic DNA to germline DNA
The chosen V,D and J genes are spliced together, with a loss of the intravening region
—-Selection is random
WHich minigene is the one transcribed to make the Ig??
The nearest minigene to the constant region is the one transcribed to make Ig
T or F
• Sometimes rearrangements don’t work. If the spliced DNA can’t be transcribed into RNA, then it hasn’t worked.
T
(blank) is also how the naïve B cell can express two different antibodies, IgM and IgD
Differential splicing
Describe the sequential events of B cell responses?
Stem cells in bone marrow make b cells-> naive or virgin make IgD or IgM-> Antigen response-> isotype switch> plasma cells form -> More B cells are made in peyers patches, spleen, lymph nodes-> antibodies in blood-> production of IgG, IgA, IgE
Rearrangments to make different antibodies sometimes doesnt work. If the spliced DNA cant be transcribed into RNA then it hasnt worked. What are the reasons that it wont work?
if both chromosomes are unable to survive rearrangment. Each B cell has 2 changes to rearrange the heavy chain and four chances to rearrange the light chain (kappa or lambda)
How can somatic mutation increase affinity?
one B cell has an antibody that binds to antigen. The affinity is low but still enough to activate the B cell. So the B cell starts to divide. TO improve the affinity, there is somatic mutation-> this process is called affinity maturation.
Within the V (D) J regions, there are hot spots of mutation. Each time the B cell divides, the hot spots of mutation (blank) the VDJ regions. What does this mean?
mutate
THis means each B cell clone is different, and so each one will have a different affinity. (some will have higher affinity, some will have worse affinity, then one with the best affinity will be selected and replicated)
Everytime the body encounters (blank), you drive more division and thus more selection for higher and higher affinity.
antigen
What is cytidine deaminase?
an inducible enzyme that can deaminate and then cause the insertion of a random nucelotide
Do T cells mutate to increase affinity?
no, they keep their original specificity
What is Ig switch controlled by?
T cell cytokine concentrations
Memory B cells have (blank), (blank), or (blank) receptors
IgA, IgG, IgE
Different types of TH pull the switches and tells the original proiferating B cell which way to go? When you have a lot of IL-4 secreted by TH2 what do your B cells make?
What if you have a lot of IFN-gamma secreted by TH1?
IgG1 and IGE
IgG3
What are polyclona antibodies?
Antibodies made in vivo
What is a monoclonal antibody (Mab)?
antibody derived in a lab
How do you make a monoclonal antibody?
prep of identical antibodies, all w/ the same L and H chains, derived from one initial immune B cell.
It is secreted by a hybrid tumor cell to form hybridoma so the cellular source of the antibody is immortal.
Can monoclonal antibodies still be cross-reactive when epitopes are similar?
Yes :(
What is Ximab?
chimeric
What zumab?
fully humanized
What is a monoclonal antibody?
a humanized monoclonal antibody is a mouse Mab to a human epitope, with the mouse constants replaced
A (blank) monoclonal antibody is a mouse Mab to a human epitope, with the mouse constants replaced. However this form still contains mouse DNA that is intervening between the genes regions that form the variable region (VDJ regions). In order to be a (blank) Mab the intervening mouse regions are replaced with human DNA. This process is repeated for the light Ig chain to complete a complete Mab
“humanized”
“fully humanized”
Monoclonal antibodies can now be made from immune human B cells
T or F
T
A human monoclonal antibody is different from a humanized mAB T or F
T
(blank) – antibody is human except for the tiny complimentarity determining region – almost no chance of rejection
humanized monoclonal antibody
