Reagents to study Wnt signaling

Reagents to study Wnt signaling can be obtained from several distributors or commercial vendors. In cases where reagents are not available, we would be happy to help , but otherwise we refer you to distributors. We have no commercial interests in those distributors or vendors, but they are professionals, can provide you with products of guaranteed quality and with the necessary background information. Reagents like purified Wnt proteins, antibodies to Wnts or to b-catenin, TOPflash reporter constructs and Wnt expression constructs can be obtained from commercial sources (R&D systems, Abcam, Cell Signaling, Upstate, or Transduction Laboratories) and easily traced by a websearch. The Wnt homepage does not include advertisements for commercial products or direct links to vendor catalogues.

This page is frequently updated. If you find anything that is published and missing on this site, alert me. If cannot find information you are looking for, then it is most likely unavailable. Specifically, beyond the Wnt-producing cells that are listed, there are no other ones available. We are trying to express other Wnts, with various degrees of success, but more work needs to be done before these are useful. Note that reagents not made in our own lab should be requested from the original sources. This is common practice in the Biomedical research community and avoids propriety issues.


cDNA clones and reporter constructs, mostly available from Addgene

Clone setcomments
Topflash, Super Topflash, other reportersFrom Randall Moon's lab
Lenti-viral vectors and reportersFrom Roel Nusse's lab (helper plasmids  pCMV∆R8.74 and  pMD2.VSVG also available from Addgene)
Wnt plasmidsFrom Jan Kitajewski'a lab
Wnt plasmidsFrom Xi He's lab
Open Source Wnt projectA great resource of cDNA clones for the Wnt genes themselves, generously provided by Drs. Marian Waterman and David Virshup. Each of the human Wnt genes has been cloned as a cDNA into expression vectors as a Gateway based library. For those who want to study mouse Wnt gene activity, be assured that the mouse/human homologs are similar enough (sometimes even identical in protein sequence) to have similar activity.

 

In general, good non-commercial sources of other cDNA clones are Addgene, the ATCC, the DGRC or the Mammalian Gene Collection, which list most of the mammalian (mouse and human) Wnt pathway cDNA clones. 


 

Wnt signaling reporter animals are listed a separate page

Many Wnt gene and Wnt pathway mutant mice, as well as "Floxed" mutant animals can be obtained from Jackson labs

 


Cells producing Active Wnt and Wingless protein (see the separate page on purifying active Wnt, which also tells you how to handle the conditioned medium).

Wnt Producing Cell

From

Name

Source

Mouse Wnt3A Nusse CRL-2647 ATCC
Mouse Wnt5A Nusse CRL-2814 ATCC
S2-Tub-wg (Drosophila Wingless S2 cells) Nusse 165 DGRC

Regular S2 cells and protocols are also available from the DGRC

Several active Wnt proteins can be purchased from R&D systems.


Antibodies (from DSHB)

Antigen

From

Name

Drosophila Frizzled 1 Adler 1C11
Drosophila Frizzled 2 Nusse 12A7
Drosophila Wingless Cohen 4D4
Drosophila Armadillo Wieschaus N2 7A1
beta-catenin Balsamo PY489-B
Ror2 NusseRor2

There are good antibodies to detect beta-catenin either on Western Blots or by staining (Transduction labs). There is also a monoclonal antibody to the non-phosphorylated ("activated") form of b-catenin (Van Noort, 2002) available from Upstate. Cell Signaling and R&D systems sell monoclonal and polyvalent antibodies to several mammalian Wnt proteins, including Wnt3A and to Wnt signaling components.

Note: Our lab is not able to send out the anti-Dishevelled or anti-Axin antibodies anymore. Because of the availability of the monoclonal antibody to Wingless (see above), we have not maintained supplies of the rabbit anti-wingless antibody and this is not available.

Fly stocks

Most of the Drosophila mutant and transgenic lines made in the Nusse lab are available through the Bloomington Stock Center, under these numbers:

38407 y[1] w[*]; wntD[KO-1] 
38408 y[1] w[*]; wntD[KO-2] 
6961UAS-DWnt2
38409 w[*]; P{w[+mC]=UASp-wntD.M}3 
38410 y[1] w[*]; P{w[+mC]=UASp-Cerk.M}2 
38411 y[1] w[*]; P{w[+mC]=UASp-Mulk.M}2/CyO 
38412 y[1] w[67c23] fz4[3-1] 
38413 y[1] w[67c23] fz4[25-1] 
38414 w[*]; P{w[+mC]=UAS-Swim.RNAi-1}3 
38415 w[*]; P{w[+mC]=UAS-Swim.M}3/TM3, Sb[1] 

Generating antibodies to Wnts, tips

It has been notoriously difficult to generate useful antibodies to vertebrate Wnt proteins. Several groups have made antibodies to peptides (Papkoff J, et al.) or Wnt-GST fusion proteins (Burrus LW, et al.). These sera work in Western blots or immunoprecipitations but usually only in extracts of cells that are engineered to over-express Wnts by transfections. In general, these sera do not detect endogenous Wnt proteins in cell extracts, nor do they detect Wnt proteins in tissues by staining techniques. Hence, there are few data on Wnt protein distribution in intact vertebrate animals.

In Drosophila however, the situation is different. There are good antisera to two Drosophila Wnt proteins: Wingless (van den Heuvel M, et al) and DWnt-3/5. (Fradkin LG, et al., Yoshikawa, 2003). It is clear why these proteins are better immunogens: they both contain long inserts in the proteins (compared to other Wnt proteins) and it has been shown that these inserts are the antigenic determinants (unpublished data, Nusse Lab). A mouse monoclonal Neumann C, et al.)to the Wingless insert works well in staining and in Western blot experiments and is available from the Developmental Studies Hybridoma Bank at The University of Iowa.

In addition, Kostriken R, et al. have made antibodies to a Leech Wnt. They took a non-conserved stretch of the protein and multimerized it to get a better immunogen.

It seems therefore that one successful strategy to get better sera is to use non-conserved parts of Wnt proteins. By generating fusion proteins (we like GST fusions) and by multimerizing the inserts, one might be able to get a useful antiserum. Note that all Wnt proteins are less conserved around the sites of the Wingless and DWnt-3/5 inserts (Alignments). These domains are also, in general, good sites for the insertion of epitopes (myc, FLAG), as we found out. Possibly, these domains stick out of the otherwise highly structurally constrained Wnt proteins and are therefore better accessible to antibodies. In addition, the Wingless and DWnt-3/5 inserts are not-conserved and are better recognized as foreign by the mammalian immune system.

Note however that because these antigenic domains in Wingless and DWnt-3/5 are not conserved, the antibodies to these proteins will not recognize any other Wnt.

Other good sites for inserting epitopes in Wnts are close to the amino-terminus.