Laboratory Members



Discover Walk for the Beckman Center

                     THE THINGS WE SEND OUT                  



HNF-1a:. An expression plasmid in pBJ5. (Courtois et al., 1987) (Baumhueter et al., 1990) (Kuo et al., 1990) (Mendel and Crabtree, 1991)
HNF-1b: An expression plasmid in pBJ5. (Courtois et al., 1987) (Baumhueter et al., 1990) (Kuo et al., 1990) (Mendel and Crabtree, 1991) (Mendel et al., 1991)
DCoH:. Transcriptional cofactor for HNF-1 binds to the amino terminus of HNF-1a and b and stablizes dimers allowing enhanced DNA binding and resistance to degradation. (Mendel et al., 1991) This was one of the first transcriptional cofactors to be cloned. The crystal structure was done in collaboration with Tom Albers lab and shows a remarkable resemblence to TBP. (Endrizzi et al., 1995)
Antibodies to HNF-1a and b are no longer available.
The alternative splice regions of the g fibrinogen genes (Crabtree and Kant, 1981) (Crabtree and Kant, 1982) are no longer available but can be obtained from the Davies Lab at the University of Washington.

The regulatory regions of the HNF-1a gene: (MODY3) that are controlled by HNF-4 (MODY1), which Calvin Kuo in the lab used to define the transcriptional cascade (Kuo et al., Nature 1992) that Graham Bell’s lab later showed was the cause of Maturity Onset Diabetes of the Young (MODY) as shown in the illustration below.

A Transcriptional Hiearchy Involved in Hepatic Development and Diabetes



Human Protein C: cDNA clones and genomic clones (Plutzky et al., 1986) are no longer available. However Jorge Plutzky may still have them. Protein C is now being marketed by Lilly as Xigeris for treatment of sepsis.

NFAT signaling
NFAT transcription complexes are assembled in the nucleus from cytoplasmic (NFATc) and nuclear (NFATn) components (Flanagan et al Nature 1991) allowing them to integrate signaling pathways (Crabtree, Science 1989). The cytoplasmic subunits are dephosphorylated by calcineurin and are exquisitely cyclosporin- and FK506-sensitive. A wide variety of signals result in Ca2+ increases, activation of calcineurin and the dephosphorylation of the different NFATc proteins. These signals include non-receptor tyrosine kinases such as the T cell receptor, most tyrosine kinase receptors, non-concanonical wnt signals, L type calcium channels, neurotropins, netrins and gap junctions. Indeed it may be that this signaling pathway is the most widely used in vertebrates. It has been difficult to detect genetically because of redundancy in the components of the pathway and is also vertebrate-specific. Biochemically, it is also difficult to detect in that most nuclear extract procedures throw out the fractions that contain NFAT transcription complexes.(see protocols) and a specialized extract procedure was originally developed to detect it (Shaw et al Science 1988).IL-2 luciferase: Originally made by David Durand and JP Shaw (Siebenlist et al., 1986). One plasmid has the 650 bp fragment from Cla1 to a synthetic HindIII site at +51 in the IL-2 gene. The other plasmid (15D Cx) has the same region truncated to 319 bp upstream of the cloned upstream of the luciferase gene (Durand et al., 1987). Both behave the same in all of the assays we have used. The mouse, human and Gibbon Ape regions (Durand et al., 1986) have all been compared and show no difference in behavior. The clone 15D Cx has been studied in transgenic mice and gives accurate expression (Emmel et al.,Science 1989)limited to activated T cells.

NF-AT luciferase: This plasmid has three copies of the NF-AT site cloned upstream of the minimal IL-2 promoter from -89 to +51 (Shaw et al., 1988) (Durand et al., 1988). This is the plasmid that was used to define NF-AT and much of the Ca2+/calcineuruin/NF-AT signaling pathway (Emmel et al., 1989) (Flanagan et al., 1991) (Clipstone and Crabtree, 1992) (Timmerman et al., 1996)

NFIL-2A Luciferase. (Durand et al., 1988) (Ullman et al., 1991)Although this plasmid binds the cJun protein and this site was used to purify JunD (Ullman et al., 1993) it behaves very similar to the NF-AT luciferase plasmid and NFAT transcription complexes with c-Jun may function at this site.
AP-1 Luciferase, Prepared from the sequences defined by Pam Mitchell in the Bob Tjian’s laboratory from the SV-40 promoter. The actual IL-2 AP-1 site is not very active in our hands.

NFkB Luciferase, Prepared from the kB enhancer as described from the Baltimore laboratory. The IL-2 NFkB site is not active in our hands, however other people have seen some activity with it.

CAML, Ca2+ modulatory Ligand of Cyclophilin. (Bram and Crabtree, 1994). This plasmid can be obtained from Rick Bram at the University of Minnesota.
CalcineurinA Prepared by Neil Clipstone from the Murine clone encoding the A1 subunit of calcineurin. (Clipstone and Crabtree, 1992). There are three genes that encode the catalytic subunit.

Calcineurin B1 Prepared by Neil Clipstone from the murine clone encoding the
B1 regulatory subunit of calcineurin. There are two genes that encode the regulatory subunit. (Clipstone and Crabtree, 1992)

The NF-ATc Genes

The nomenclature is confusing for these proteins. The one below is the original one (Flanagan et al Nature 1991 and Crabtree and Scbreiber 1992). It is the offical nomenclature and can be found at the Genome Database site: To find the new nomenclature look for: The cytoplamic, cyclosporin-senstive subunits are encoded by 4 genes. The nuclear subunits have not yet been purified but appear to be AP-1 and other proteins.

NF-ATc1(c), Expression vector for NF-ATc1 in a pBJ5 plasmid. The monoclonal antibodies that we made to NF-ATc1 are sold by Pharmagen and Santa Cruz. (Northrop et al., 1994). The crystal structure revealed that our initial predictions that it was a rel-like protein were in fact correct. The DNA binding domain is defective compared to a classic DNA binding domain (Wolfe et al., 1997) hence allowing the assembly of NFAT transcription complexes to integrate signals.

NF-ATc2(p), Expression vector for human NF-ATc2 in a pBJ5 plasmid (Northrop et al., Nature 1994). Sequence for the human and bovin clones were obtained by Jeff Northrop and Steffan Ho.

NF-ATc3(x)(4), Expression vector for NF-ATc3 in a pBJ5 plasmid. (Ho et al., 1995). In this publication a full length protein was cloned, but it contains a start-site that is not present in most of the other isolates, hence we thought it was not full lenght,but it actually is.

NF-ATc4(3), Expression vector for murine NF-ATc4 in a pBJ5 plasmid (Greaf et al Nature 1999, Greaf et al Cell 2001 and Greaf et al Cell 2003).

TonEBP (NF-AT5)- Originally discovered by cloned by Kron and colleagues at John Hopkins. A cDNA was isolated by Steffan Ho, who now has these clones (see past lab members for address)

Prolyl Isomerases and Immunophilins

Cyclophilins A to D (Bram et al., 1993) These are expression vectors used by Rick Bram to define the cyclophilins that were active in blocking the actions of calcineurin.

FKBP12, 13, 25 and 52 were obtained from Stuart Schreiber’s laboratory

CRAC-Channel Deficient Cell Lines. These were isolated by Andrew Serafini in the lab(Serafini et al., 1995). The mutations appear to be dominant and have not yet been cloned. (Clipstone and Crabtree, 1992)

TAG Jurkats- This cell line was created by Steve Fiering and Jeff Northrop in the lab and has SV-40 T antigen and a integrated NF-AT b gal plasmid (Fiering et al., 1990), (Clipstone and Crabtree, 1992).


Yeast Genes Involved in the Rapamycin-Sensitive Growth Regulatory Pathway.

: (Zheng et al., 1995) (Fiorentino and Crabtree, 1997). Identical to the clones isolated by M. Hall.
TOR2: (Zheng et al., 1995) (Fiorentino and Crabtree, 1997). Identical to the clones isolated by M. Hall.
DNA-2 (ROT1) (Fiorentino and Crabtree, 1997)We have given our clones and ts allels to Judith Campbell to distribute

Components of SWI/SNF-Like BAF Complexes

BAF complex

The names used are the ones that we originally used in characterizing the complex, however since neither SWI/SNF or BAF was acceptable to the Genome Nomenclature Committee we renamed them as "Swi-Related, Matrix-associated, Actin Dependent Regulators of Chromatin or SMARC protiens. To find the new nomenclature look for:

BAF250a, b,and c (SMARC d1, d2, Arid2) These can be obtained from Weidong Wang now a senior investigator at the National Institutes of Health. BAF250c is also called Arid or BAF200.

BAF 190a (BRG1) (SMARCA4) (Khavari et al., 1993) (Dunaief et al., 1994) (Zhao et al., 1998) This plasmid is an expression vector for the full length BRG/BAF190 protein which is similar to Drosphila Brm and contains the conserved ATPase domain in the yeast SWI2/SNF2 protein. We also have a dominant negative (Khavari et al Nature 1993), which we distributed widely, but now know is a gain-of-function since it gives a phenotype that is more severe than the knock out of both Brg and Brm. Hence, we no longer recommend that the dominant negative be used to analyze the function of this complex.

HBrm- BAF190b (SMARCA2)
This clone was isolated in Mose Yanif’s lab at the Pasteur Insitute and is distributed by him. It is highly homologous to Brg, but Brm complexes appear to have a different function than Brg complexes based on the phenotypes of mice with mutations in these proteins.
Anti BRG1, JI is a rabbit antibody to the C terminus of BRG1. It is good for most everything you need to do with this complex (Khavari et al., 1993) (Dunaief et al., 1994) (Zhao et al., 1998).

BAF 170 (SMARCC2), This is an expression plasmid based on pBJ5. BAF170 has a domain related to SWI3. (Wang et al., 1996)

BAF 155 (SMARCC1, This is an expression plasmid based on pBJ5.BAF155 has a domain related to SWI3. (Wang et al., 1996)


BAF 60a, (SMARCD1) (Wang et al., 1996) This is an expression plasmid based on pBJ5. BAF60a is related to a component of the yeast SWI/SNF complex found by Brad Crains and reported in Genes and Development 1996.

BAF 60b
(SMARCD2)(Wang et al., 1996). This is an expression plasmid based on pBJ5. BAF60a is related to a component of the yeast SWI/SNF complex found by Brad Crains and reported in Genes and Development 1996.

BAF 60c (SMARCD3)(Wang et al., 1996). This is an expression plasmid based on pBJ5. BAF60a is related to a component of the yeast SWI/SNF complex found by Brad Crains and reported in Genes and Development 1996.

(Wang et al., 1998). BAF57 has a kinesin coiled-coil domain and a HMG domain. There is no homologue of this protein in the yeast SWI/SNF complex. BAF57 (SMARCE1) antibodies made in rabbits are also available. These IP as well as Western blot and are also useful for immunofluorescene and CHIP assays. See Chi et al Nature 2002

BAF 53a (Actl6b) (Zhao et al., 1998). This is an actin related member of of the ARP family of proteins. It directly interacts with BRG1 in the complex. Shortly after we discovered this family member both Braid Crains and Craig Peterson found yeast homologues in the yeast data base called ARP7 and ARP9 that are components of the yeast SWI complex. The yeast homologues are less related to actin than BAF53a. Hence we do not yet know if there is going to be a functional relationship between the yeast proteins and the mammalian proteins.

BAF 53b (Actl6a) This subunit is neuron specific and has not been found to date in other cell types. It is highly related to BAF53a.
BAF 47 (Kalpana et al., 1994) Also called INI for integrase interacting protein or hSNF5 because it shares a domain with yeast SNF5 isolated in Marion Carlson’s laboratory.

b-Actin. (Zhao et al., 1998). This is a component of the mammalian complex but not the yeast complex. We feel that it is the most distinctive feature of the mammalian BAF complex and probably indicates that BAF is not likely to be mechanistically related to the yeast complex. We have b-actin with mutations in the nuclear export sequence and also GFP tagged versions of the protein.


Brd7 A newly discovered subunit not yet found in yeast or flies.


Brd9 A newly discovered subunit essential for the activity of the complex in es cells.












  Design and Content by G R Crabtree