Resources

CD40 and B-cell receptor signalling induce MAPK family members that can either induce or repress Bcl-6 expression

Cross-linking surface IgM with goat anti-IgM.

http://www.sciencedirect.com/science/article/pii/S0161589009000704

Fusion protein linkers: property, design and functionality

Gives examples of fusion protein linkers - recommend (GGGGS)₃ for flexible linker

http://www.sciencedirect.com/science/article/pii/S0169409X12003006

Early BCR events and antigen capture, processing, and loading on MHC class II on B cells

Overview of early BCR activation events (see BCR Signaling and Antigen Internalization). Paper by Ana Avalos, the person that Jas was going to put us in contact with.

http://journal.frontiersin.org/Journal/10.3389/fimmu.2014.00092/full

Src-family kinases in B-cell development and signaling

Review about BCRs.  Specifically contains useful information about Lyn and how Lyn and Syk activation work.

http://www.nature.com/onc/journal/v23/n48/full/1208075a.html

More on the BCR signalling pathway:

http://www.annualreviews.org/doi/full/10.1146/annurev.immunol.17.1.555

Syk does seem to be recruited to the receptor upon binding (at least in B cells):

http://www.jimmunol.org/content/166/3/1507.long

This also shows that Syk fusion proteins seem to still be functional (at least when it's stuck to the C terminus).

According to this, SykK396R point mutation is catalytically inactive; however, recruitment is less efficient.

Syk does a LOT of stuff:

http://www.nature.com/nri/journal/v10/n6/full/nri2765.html

If the cell already expresses Syk, maybe we could knock out the domain of Lyn that phosphorylates it?

Thread on signal sequences we can use for our antibody chains

http://www.researchgate.net/post/Can_anyone_suggest_a_leader_peptide_sequence_for_efficient_secretion_of_a_recombinant_protein_transiently_expressed_in_293_cells

Are we sure the light-chain signal peptide is the part we think it is?

It seems to be - http://www.nature.com/nature/journal/v288/n5792/pdf/288730a0.pdf

What are all the different possible signal peptide sequences?

List here (of "leader regions") - http://www2.mrc-lmb.cam.ac.uk/vbase/alignments2.php

There may be a few that aren't on this list, though. (for example VH1-69 which was from the example sequence we found doesn't seem to be on here.)

Is there a point mutation that makes Lyn constitutively phosphorylated?

There seems to be - mutating Tyr508, changing it to F, blocks inhibition of autophosphorylation/activation.

http://www.jbc.org/content/281/42/31920.long

This makes Lyn constitutively active.

http://jem.rupress.org/content/196/12/1593.full

Where is Lyn normally localized?

It seems to get stuck to membrane proteins anyway - http://jcb.rupress.org/content/165/5/641.full

Relevant to Syk

Lyn phosphorylates Syk at Tyr323

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2746503/ (Syk structure)

also rather syk: http://www.sciencedirect.com/science/article/pii/S0022283612008790

It is possible that Syk is associated with the complex before activation (although these papers are relatively old, so this may have been disproved since then) -

http://www.jbc.org/content/267/12/8613.long

seems relevant to lyn phosphorylation:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC359265/pdf/molcellb00011-0280.pdf

annotation for a kappa-chain variable region:

http://www.uniprot.org/uniprot/P01602

(the signal sequence here is MDMRVPAQLLGLLLLWLPGAKC)

TRANSFECTED PLASMACYTOMA CELLS DO NOT TRANSPORT THE MEMBRANE FORM OF IgM TO THE CELL SURFACE

cDNA can produce IgM that localizes to the membrane.  Also includes information on using FACS to test for surface IgM expression.

http://jem.rupress.org/content/167/2/652.long

Structure of the 5′ Ends of Immunoglobulin Genes: A Novel Conserved Sequence

The initial signal sequences of kappa light chain variable regions are variable, 16-26 amino acid sequences (p. 2651).

http://www.jstor.org.libproxy.mit.edu/stable/23391

B lymphocyte antigen receptor signaling: initiation, amplification, and regulation

A 2013 review of current understanding and controversy over BCR mechanisms.

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3790562/

Transfecting IgM, CD79A, and CD79B is sufficient to yield BCR surface expression:

http://www.pnas.org/content/89/8/3404.long