Key to Organismal Survival and Continued Propagation Is Maintenance of Genome Integrity

Cell line BRG1 p53 Reference Hela + WT [1] U2OS + WT [2] MCF7 + WT [2] HepG2 + WT [3] Y79 + WT [3] HEK293 + WT [2] G401 + WT [3] RKO + WT [4] & This study LNCaP + WT [5] SW13 deficient mut [6, 7] ALAB null null [8] H1299 null null [9] C33a deficient mut [2, 10] PANC1 deficient mut [3] TSU-Pr1 deficient mut [11] DU145 deficient mut [9] Jurkat + mut [3]

Supplementary Table 1: List of tumor derived cell lines indicating p53 genotype and BRG1 expression status. + = Detectable level of expression by western, deficient = levels can not be detected by western and or do not express transcript.

1. Sif, S., et al., Mitotic inactivation of a human SWI/SNF chromatin remodeling complex. Genes Dev, 1998. 12(18): p. 2842-51. 2. Strobeck, M.W., et al., BRG-1 is required for RB-mediated cell cycle arrest. Proc Natl Acad Sci U S A, 2000. 97(14): p. 7748-53. 3. Decristofaro, M.F., et al., Characterization of SWI/SNF protein expression in human breast cancer cell lines and other malignancies. J Cell Physiol, 2001. 186(1): p. 136-45. 4. Kuerbitz, S.J., et al., Wild-type p53 is a cell cycle checkpoint determinant following irradiation. Proc Natl Acad Sci U S A, 1992. 89(16): p. 7491-5. 5. Wang, X., et al., Expression of p270 (ARID1A), a component of human SWI/SNF complexes, in human tumors. Int J Cancer, 2004. 112(4): p. 636. 6. Muchardt, C. and M. Yaniv, A human homologue of Saccharomyces cerevisiae SNF2/SWI2 and Drosophila brm genes potentiates transcriptional activation by the glucocorticoid receptor. Embo J, 1993. 12(11): p. 4279-90. 7. Dunaief, J.L., et al., The retinoblastoma protein and BRG1 form a complex and cooperate to induce cell cycle arrest. Cell, 1994. 79(1): p. 119-30. 8. Hendricks, K.B., F. Shanahan, and E. Lees, Role for BRG1 in cell cycle control and tumor suppression. Mol Cell Biol, 2004. 24(1): p. 362-76. 9. Wong, A.K., et al., BRG1, a component of the SWI-SNF complex, is mutated in multiple human tumor cell lines. Cancer Res, 2000. 60(21): p. 6171-7.

10. Zhang, H.S., et al., Exit from G1 and S phase of the cell cycle is regulated by repressor complexes containing HDAC-Rb-hSWI/SNF and Rb-hSWI/SNF. Cell, 2000. 101(1): p. 79- 89. 11. van Bokhoven, A., et al., TSU-Pr1 and JCA-1 cells are derivatives of T24 bladder carcinoma cells and are not of prostatic origin. Cancer Res, 2001. 61(17): p. 6340-4.

Supplemental Figure S1. SWI/SNF subunit Brm is not required for p53 degradation.

Upper panel immunoblots of HeLa cell extracts of control and shBrm plasmids transfected and blotted with antibodies as indicated. Lower panel immunoblots of HeLa cells extracts transfected with control or shBrm and treated with cycloheximide and harvested after indicated time points and blotted with antibodies to p53 and actin. CHX = cycloheximide

Supplemental Figure S2. RNAi knockdown of BRG1 blocks p53 poly-ubiquitination.

Top panel: U2OS cells transfected with control or shBRG1 plasmids and immunoblotted for endogenous p53, BRG1 and actin. Bottom panel: U2OS cells transfected with control or BRG1 shRNA and cell extracts immunoprecipitated with IgG or p53 antibodies followed by immunoblotting with antibodies to ubiquitin. 5% input levels of p53 and actin from these cell extracts are shown below.

Supplemental Figure S3. p21 activation due to BRG1 depletion is p53 dependent. Upper left panel: control and R248W dominant negative (dn) p53 expressing HeLa cells were transfected with control or BRG1 shRNA and the extracts were immunoblotted for p53, BRG1, p21 and actin. Upper right panel: control and dnp53 HeLa cells transfected with control or BRG1 shRNA were stained for SA ß-gal. 500 cells were scored and % positive cells plotted. Bottom panel: Same as upper right panel showing representative fields and ß-gal positive cells.

Supplemental Figure S4. NBRG, the BRG1 deletion of the PRR, complexes with

BAF47 and stimulates transcription.

(A) H1299 cells transfected with Flag-NBRG or Flag-BRG1 were immunoprecipitated with IgG or Flag antibodies and immunoblotted for endogenous BAF47 and Flag-tagged proteins. (B) SW13 cells transfected with vector control, NBRG or BRG1 and levels of

CD44 and GAPDH transcripts were evaluated by RT-PCR.

Supplemental Figure S5. BRG1 mediated p53 inactivation requires the proline rich region (PRR). C33a cells transfected with indicated plasmids, selected with puromycin for 3 weeks, stained with crystal violet, and colonies counted.

Supplemental Figure S6. BRG1 depletion does not affect p21 expression in p53-null PC3 cells. Total extracts of PC3 cells transfected with control or shBRG1 were blotted for BRG1, p21 and actin.

Supplemental Figure S7. BRG1 depletion does not change the expression levels of E6AP and E6 proteins in HeLa cells. HeLa cell extracts prepared from control or BRG1 shRNAs were blotted for E6AP, E6 (monoclonal antibody C1P5) and actin.