Apaf-1 is a Transcriptional Target for the ZNF217 Oncogene

Institution: University of California, Davis
Investigator(s): Sheryl Krig, Ph.D. -
Award Cycle: 2005 (Cycle 11) Grant #: 11FB-0029 Award: $53,649
Award Type: Postdoctoral Fellowship
Research Priorities
Biology of the Breast Cell>Pathogenesis: understanding the disease

Initial Award Abstract (2005)
The ZNF217 oncogene is amplified and over-expressed in 20-30% of early stage breast tumors, and it appears to play a key role during early transformation of normal human mammary epithelial cells (HMEC). Research with ZNF217 has shown that inserting this oncogene into HMEC will allow them to grow indefinitely and immortalize. In addition, preliminary data from our lab and collaborators, suggests ZNF217 protects breast tumor cells from cell death upon doxorubicin treatment. Such chemotherapeutic agents, used in advanced stages of cancer, cause DNA double-stranded breaks that trigger cell death pathways leading to elimination of the tumor cell. We hypothesize that ZNF217 prevents the execution of the cell death pathways, both early in the immortalization process and later, at malignant stages of tumor progression, allowing abnormal cells to survive.

Based on its protein structure, it is likely that ZNF217 is a DNA-binding protein and acts at specific DNA sites in promoter regions, along with other proteins, in a transcription regulatory complex. Our recent data suggests ZNF217 binds to and represses the Apaf-1 (apoptotic protease-activating factor-1) promoter, which would otherwise trigger cell death pathways. We are now performing additional analyses using chromatin immunoprecipitation (ChIP) assays to identify other promoters regulated by ZNF217. We are also characterizing ZNF217-regulated promoters to determine if ZNF217 contributes to neoplastic transformation by causing changes in chromosomal structure. Identifying ZNF217 target genes and characterizing how ZNF217 functions to regulate genes such as Apaf-1 will facilitate the design of drugs to make chemotherapy more effective.

Final Report (2006)
My studies of ZNF217 over the past 15 months have provided important leads in understanding how aberrant expression of this putative transcriptional regulator promotes the transformation of a normal cell into a tumor cell. At the start of this project, I had hypothesized that ZNF217 either directly or indirectly binds DNA and is associated with a transcription regulatory complex at specific promoter regions. To test this hypothesis, I used an unbiased global approach (chromatin immunoprecipitation followed by microarray analysis) to identify promoters regulated by ZNF217.

Using this "ChIP-chip" method I have identified target genes for ZNF217 in three different cell lines; the MCF7 breast cancer cell line, the SW480 colon cancer line, and the Ntera2 embryonal teratocarcinoma line. I have established that ZNF217 targets genes in a cell-type specific manner. Using siZNF217-treated (i.e., using inhibitory RNA to reduce ZNF217 expression) Ntera2 cells, I have probed expression arrays to determine whether ZNF217 activates or represses the specific targets that it binds to. Bioinformatics analysis was used to identify a motif (protein sub-region) that is involved in recruiting ZNF217 to the chromatin. By “gene ontology” (see this reference for a description of this term) analysis of the set of ZNF217 target genes, I have identified enrichment for transcription factor activity and developmental pathways. Further work with Ntera2 cells shows that retinoic acid differentiation leads to down-regulation of ZNF217 levels by Day 2 of treatment.

We hypothesize that the normal function of ZNF217 may be to suppress differentiation-specific genes and help maintain a proliferative state. Future analysis of retinoic acid-induced differentiation of stem cells may demonstrate pathways that are affected by aberrant ZNF217 expression in tumors, leading to insight into the role of ZNF217 in the immortalization process and tumor development.

Suz12 binds to silenced regions of the genome in a cell-type-specific manner.
Periodical:Genome Research
Index Medicus: Genome Res
Authors: Squazzo SL, O'Geen H, Komashko VM, Krig SR, et al.
Yr: 2006 Vol: 16 Nbr: 7 Abs: Pg:890-900

Identification of genes directly regulated by the oncogene ZNF217 using chromatin immunoprecipitation (ChIP)-chip assays.
Periodical:Journal of Biological Chemistry
Index Medicus: J Biol Chem
Authors: Krig SR, Jin VX, Bieda MC, O'Geen H, Yaswen P, Green R, Farnham PJ.
Yr: 2007 Vol: 282 Nbr: 13 Abs: Pg:9703-9712