Syk-dependent alternative homologous recombination activation promotes cancer resistance to DNA targeted therapy.

Enhanced DNA repair is an important mechanism of inherent and acquired resistance to DNA targeted therapies, including poly ADP ribose polymerase inhibition. Spleen associated tyrosine kinase (Syk) is a non-receptor tyrosine kinase known to regulate immune cell function, cell adhesion, and vascular development. Here, we report that Syk can be expressed in high grade serous ovarian cancer and triple negative breast cancers and promotes DNA double strand break resection, homologous recombination (HR) and therapeutic resistance. We found that Syk is activated by ATM following DNA damage and is recruited to DNA double strand breaks by NBS1. Once at the break site, Syk phosphorylates CtIP, a key mediator of resection and HR, at Thr-847 to promote repair activity, specifically in Syk expressing cancer cells. Syk inhibition or genetic deletion abolished CtIP Thr-847 phosphorylation and overcame the resistant phenotype. Collectively, our findings suggest that Syk drives therapeutic resistance by promoting DNA resection and HR through a novel ATM-Syk-CtIP pathway, and that Syk is a new tumor-specific target to sensitize Syk-expressing tumors to PARPi and other DNA targeted therapy.

Identification and characterization of the human SCAN domain zinc-finger gene ZNF449.

Zinc-finger proteins play important roles in various cellular functions, including cell proliferation, differentiation, and apoptosis. In this study, we identified a gene encoding a zinc finger protein named ZNF449, which was isolated from human testis cDNA library and mapped to Xq26.3 by searching the UCSC genomic database. The ZNF449 cDNA consists of 4035 nucleotides and has a 518-amino acids open reading frame. The predicted protein contains a leucine-rich region (LER or SCAN domain) at the N-terminus. At the C-terminus of the protein, there are 7 C(2)H(2) (Cys2-His2) zinc-finger motifs. The expression pattern of ZNF449 showed that it was ubiquitously expressed in 18 human adult tissues. ZNF449 protein was located in the nucleus when overexpressed in cultured cells. These results indicate that ZNF449 is a member of the zinc-finger family and it may function as a transcription factor.

Activation of transcriptional activities of AP1 and SRE by a novel zinc finger protein ZNF445.

Zinc finger proteins play important roles in various cellular functions, including cell proliferation, differentiation, and apoptosis. Mitogen-activated protein kinase (MAPK) signal transduction pathways are one of the most common mechanisms in eukaryotic cell regulation. Many transcription factors are important targets of MAPKs. In this study, we identified a novel gene encoding a zinc finger protein named ZNF445. The ZNF445 mRNA consists of 9105 nucleotides and has a 1031-amino acid open reading frame. The predicted 119-kDa protein contains a leucine-rich region (LER or SCAN domain) at the N-terminus, followed by a well-conserved Krüppel-associated box (KRAB) domain. At the C-terminus of the protein, there are 14 C2H2 (Cys2-His2) zinc finger motifs. ZNF445 gene is mapped to chromosome 3p21.32. Northern blot analysis indicates that a 9.1 kb transcript specific for ZNF445 is expressed in uterus, thymus, small intestine, colon, pancreas, peripheral blood leukocyte, and especially at a higher level in the testis and skeletal muscle in human adult tissues. ZNF445 protein was located in the nucleus when overexpressed in cultured cells. Reporter gene assays showed that ZNF445 is a transcriptional repressor, and overexpression of ZNF445 in the HEK 293T cells activates the transcriptional activities of AP1 and SRE. Deletion studies showed that the SCAN domain of ZNF445 may be involved in this activation. Furthermore, we found that expression of ZNF445 can increase p42/44 MAPK, MEK and Raf-1 phosphorylation. These results clearly indicate that ZNF445 is a member of the zinc finger transcription factor family and may function in MAPK pathway through Raf-1/MEK/p42/44 MAPK signals.

Cloning and characterization of the CDZFP gene which encodes a putative zinc finger protein.

We report here the cloning and characterization of a novel human cytoplasm-distribution zinc finger protein (CDZFP) gene, isolated from human ovary cDNA library, and mapped to 4p12 by searching the UCSC genomic database. The CDZFP cDNA is 1793 base pairs in length and contains an open reading frame (ORF) encoding 236 amino acids. The CDZFP gene consists of 7 exons and encodes a putative zinc finger protein with a transmembrane region and two zinc finger motifs. Subcellular localization demonstrated that CDZFP protein was located in the cytoplasm when overexpressed in Hela cells and northern blot analysis revealed that CDZFP was ubiquitously expressed in 16 human tissues.

Identification and characterization of two novel human SCAN domain-containing zinc finger genes ZNF396 and ZNF397.

We have identified two novel human SCAN domain genes ZNF396 and ZNF397, which are clustered within the region of chromosome 18q12. Three isoforms of ZNF396 transcript, 1.5, 2.5 and 3.9-kb, are expressed highly in liver. Four isoforms of ZNF397 transcript, 1.7, 2.5, 7.0 and 9.0-kb, are expressed in a variety of tissues, with varying levels. The SCAN-(C(2)H(2))(X) genes encode two distinct proteins due to a unique alternative splicing mechanism. Both ZNF396-fu (full zinc fingers) and ZNF397-fu consist of a SCAN domain in the N-terminal region and many consecutive C(2)H(2) zinc finger repeats in the C-terminal region. ZNF396-nf (no zinc fingers) and ZNF397-nf encode 210 and 198 amino acids, respectively, containing the SCAN domain only. ZNF396-fu, ZNF396-nf, ZNF397-fu or ZNF397-nf can homo-associate, while ZNF396-fu hetero-associates with ZNF396-nf, and ZNF397-fu hetero-associates with ZNF397-nf. ZNF396-nf and ZNF397-nf polypeptides are expressed diffusely in the cells, while ZNF396-fu and ZNF397-fu polypeptides target specifically to the nuclei. ZNF396-fu, ZNF396-nf and ZNF397-nf can repress reporter gene transcription, with ZNF397-nf having the strongest repression activity. Deletion analysis revealed that ZNF397-fu is a transcriptional activator without its nine zinc finger repeats.