SCAN Box Research Articles

Context-dependent pro- and anti-resection roles of ZKSCAN3 in the regulation of fork processing during replication stress

Uncontrolled resection of replication forks under stress can cause genomic instability and influence cancer formation. Extensive fork resection has also been implicated in the chemosensitivity of “BReast CAncer gene” BRCA-deficient cancers. However, how fork resection is controlled in different genetic contexts and how it affects chromosomal stability and cell survival remains incompletely understood. Here, we report a novel function of the transcription repressor ZKSCAN3 in fork protection and chromosomal stability maintenance under replication stress. We show disruption of ZKSCAN3 function causes excessive resection of replication forks by the exonuclease Exo1 and homologous DNA recombination/repair protein Mre11 following fork reversal. Interestingly, in BRCA1-deficient cells, we found ZKSCAN3 actually promotes fork resection upon replication stress. We demonstrate these anti- and pro-resection roles of ZKSCAN3, consisting of a SCAN box, Kruppel-associated box, and zinc finger domain, are mediated by its SCAN box domain and do not require the Kruppel-associated box or zinc finger domains, suggesting that the transcriptional function of ZKSCAN3 is not involved. Furthermore, despite the severe impact on fork structure and chromosomal stability, depletion of ZKSCAN3 did not affect the short-term survival of BRCA1-proficient or BRCA1-deficient cells after treatment with cancer drugs hydroxyurea, PARPi, or cisplatin. Our findings reveal a unique relationship between ZKSCAN3 and BRCA1 in fork protection and add to our understanding of the relationships between replication fork protection, chromosomal instability, and chemosensitivity.

Effects of remote digital monitoring on oral hygiene of orthodontic patients: a prospective study

BackgroundRemote digital monitoring during orthodontic treatment can help patients in improving their oral hygiene performance and reducing the number of appointments due to emergency reasons, especially in time of COVID-19 pandemic where non-urgent appointments might be discouraged.MethodsThirty patients scheduled to start an orthodontic treatment were divided into two groups of fifteen. Compared to controls, study group patients were provided with scan box and cheek retractor (Dental Monitoring®) and were instructed to take monthly intra-oral scans. Plaque Index (PI), Gingival Index (GI), and White Spot Lesions (WSL) were recorded for both groups at baseline (t0), every month for the first 3 months (t1, t2, t3), and at 6 months (t4). Carious Lesions Onset (CLO) and Emergency Appointments (EA) were also recorded during the observation period. Inter-group differences were assessed with Student's t test and Chi-square test, intra-group differences were assessed with Cochran’s Q-test (significance α = 0.05).ResultsStudy group patients showed a significant improvement in plaque control at t3 (p = 0.010) and t4 (p = 0.039), compared to control group. No significant difference was observed in the number of WSL between the two groups. No cavities were detected in the study group, while five CLO were diagnosed in the control group (p = 0.049). A decreased number of EA was observed in the study group, but the difference was not significant.ConclusionsIntegration of a remote monitoring system during orthodontic treatment was effective in improving plaque control and reducing carious lesions onset. The present findings encourage orthodontists to consider this technology to help maintaining optimal oral health of patients, especially in times of health emergency crisis.

Macaca specific exon creation event generates a novel ZKSCAN5 transcript

ZKSCAN5 (also known as ZFP95) is a zinc-finger protein belonging to the Krűppel family. ZKSCAN5 contains a SCAN box and a KRAB A domain and is proposed to play a distinct role during spermatogenesis. In humans, alternatively spliced ZKSCAN5 transcripts with different 5′-untranslated regions (UTRs) have been identified. However, investigation of our Macaca UniGene Database revealed novel alternative ZKSCAN5 transcripts that arose due to an exon creation event. Therefore, in this study, we identified the full-length sequences of ZKSCAN5 and its alternative transcripts in Macaca spp. Additionally, we investigated different nonhuman primate sequences to elucidate the molecular mechanism underlying the exon creation event. We analyzed the evolutionary features of the ZKSCAN5 transcripts by reverse transcription polymerase chain reaction (RT-PCR) and genomic PCR, and by sequencing various nonhuman primate DNA and RNA samples. The exon-created transcript was only detected in the Macaca lineage (crab-eating monkey and rhesus monkey). Full-length sequence analysis by rapid amplification of cDNA ends (RACE) identified ten full-length transcripts and four functional isoforms of ZKSCAN5. Protein sequence analyses revealed the presence of two groups of isoforms that arose because of differences in start-codon usage. Together, our results demonstrate that there has been specific selection for a discrete set of ZKSCAN5 variants in the Macaca lineage. Furthermore, study of this locus (and perhaps others) in Macaca spp. might facilitate our understanding of the evolutionary pressures that have shaped the mechanism of exon creation in primates.

MAGE proteins regulate KRAB zinc finger transcription factors and KAP1 E3 ligase activity

Expression of Melanoma AntiGen Encoding (MAGE) genes, particularly MAGE-A3, has been correlated with aggressive clinical course, the acquisition of resistance to chemotherapy and poor clinical outcomes of melanoma and other malignancies. MAGE proteins bind to KAP1, a gene repressor and ubiquitin E3 ligase which also binds KRAB domain containing zinc finger transcription factors (KZNFs), and MAGE expression may affect KZNF mediated gene regulation. To investigate mechanisms for these effects, we tested the hypothesis that differences in KRAB domain composition affect KZNF poly-ubiquitination and determine whether MAGE expression increases, decreases, or has no effect on KZNFs mediated gene repression. Using an integrated reporter gene responsive to repression by KRAB domain fusion proteins, we found that MAGE-A3 relieved KZNF mediated repression and induced KZNF poly-ubiquitination and degradation in association with expression of the A+B box KRAB domain. In contrast, MAGE-A3 enhanced KAP1 mediated repression of KZNFs expressing A or A+b box KRAB domains but caused no increase in poly-ubiquitination or degradation. MAGE-A3 has no significant impact on KZNFs with KRAB domains containing the Scan box motif. These data support our hypothesis by showing that the effects of MAGE-A3 on gene repression depend on the type of KZNF KRAB domain involved.

Measuring Diffusion Coefficients via Two-photon Fluorescence Recovery After Photobleaching

Multi-fluorescence recovery after photobleaching is a microscopy technique used to measure the diffusion coefficient (or analogous transport parameters) of macromolecules, and can be applied to both in vitro and in vivo biological systems. Multi-fluorescence recovery after photobleaching is performed by photobleaching a region of interest within a fluorescent sample using an intense laser flash, then attenuating the beam and monitoring the fluorescence as still-fluorescent molecules from outside the region of interest diffuse in to replace the photobleached molecules. We will begin our demonstration by aligning the laser beam through the Pockels Cell (laser modulator) and along the optical path through the laser scan box and objective lens to the sample. For simplicity, we will use a sample of aqueous fluorescent dye. We will then determine the proper experimental parameters for our sample including, monitor and bleaching powers, bleach duration, bin widths (for photon counting), and fluorescence recovery time. Next, we will describe the procedure for taking recovery curves, a process that can be largely automated via LabVIEW (National Instruments, Austin, TX) for enhanced throughput. Finally, the diffusion coefficient is determined by fitting the recovery data to the appropriate mathematical model using a least-squares fitting algorithm, readily programmable using software such as MATLAB (The Mathworks, Natick, MA).

Measuring Diffusion Coefficients via Two-photon Fluorescence Recovery After Photobleaching

Multi-fluorescence recovery after photobleaching is a microscopy technique used to measure the diffusion coefficient (or analogous transport parameters) of macromolecules, and can be applied to both in vitro and in vivo biological systems. Multi-fluorescence recovery after photobleaching is performed by photobleaching a region of interest within a fluorescent sample using an intense laser flash, then attenuating the beam and monitoring the fluorescence as still-fluorescent molecules from outside the region of interest diffuse in to replace the photobleached molecules. We will begin our demonstration by aligning the laser beam through the Pockels Cell (laser modulator) and along the optical path through the laser scan box and objective lens to the sample. For simplicity, we will use a sample of aqueous fluorescent dye. We will then determine the proper experimental parameters for our sample including, monitor and bleaching powers, bleach duration, bin widths (for photon counting), and fluorescence recovery time. Next, we will describe the procedure for taking recovery curves, a process that can be largely automated via LabVIEW (National Instruments, Austin, TX) for enhanced throughput. Finally, the diffusion coefficient is determined by fitting the recovery data to the appropriate mathematical model using a least-squares fitting algorithm, readily programmable using software such as MATLAB (The Mathworks, Natick, MA).

Spectroscopic characterization of the tumor antigen NY-REN-21 and identification of heterodimer formation with SCAND1

Human NY-REN-21 is a C 2H 2 type multi-finger protein, with a SCAN domain in the N-terminal region and a predicted coil central region. It represents a putative ortholog of mouse ZFP38, a transcriptional factor that recognizes a bipartite DNA motif and is unable to form homodimers. As shown in this work, NY-REN-21 contains a SCAN domain able to form homodimers and a central region that behaves as an intrinsically disordered protein. The SCAN domain is found in 71 human proteins and its ability to form homo- and heterodimers widens the number of genes that are regulated by this group of transcription factors. NY-REN-21 interaction with SCAND1 was identified using the yeast two-hybrid system and confirmed using recombinant proteins. SCAND1 is a truncated SCAN box protein, lacking the zinc finger region and the NY-REN-21/SCAND1 heterodimer is asymmetric concerning the DNA binding region. This result indicates that NY-REN-21 can function either as a homodimer or as a heterodimer with SCAND1.

Hypertrophic osteoarthropathy from pulmonary metastatic phyllodes tumour of the breast

The Medical Journal of Australia ISSN: 0025729X 6 September 2004 181 5 279-279 ©The Medical Journal of Australia 2004 www.mja.com.au Snapshots Hypertrophic osteoarthropathy (HOA) secondary to metastatic phyllodes tumour was diagnosed. HOA is a tic syndrome characterised by digital clubbing, polya periostosis. The joint pain, resistant to non-opiate analgesia, responded to intravenous pamidronate. T tumour response to chemotherapy with adriamycin and Phyllodes tumours are fibroepithelial lesions arising A 28-year-old woman presented with a 3-month history of wrist and ankle pain, lethargy and weight loss. She had clubbing of the fingers (Box 1) and toes. A bone scan (Box 2) and chest x-ray (Box 3) revealed periostosis in the wrists and pulmonary metastases, respectively. Two years earlier she had been diagnosed with a malignant phyllodes tumour of the left breast (Box 44), which was treated by mastectomy and axillary dissection. pulmonary paraneoplasrthralgia and and opiate here was no ifosfamide. from intralobular breast stroma. They account for less than 1% of breast neoplasms. Treatment is by surgical resection. About 20% of patients with phyllodes tumours develop distant metastases, and long-term survival in these women is rare. Fiona J Collinson,* A Michael Bilous,† Richard F Kefford‡ * Advanced Trainee, † Director of Tissue Pathology ‡ Professor of Medicine, Department of Medical Oncology University of Sydney at Westmead Hospital, Westmead, NSW fjcollinson@doctors.org.uk

Nizp1, a novel multitype zinc finger protein that interacts with the NSD1 histone lysine methyltransferase through a unique C2HR motif.

Haploinsufficiency of the NSD1 gene is a hallmark of Sotos syndrome, and rearrangements of this gene by translocation can cause acute myeloid leukemia. The NSD1 gene product is a SET-domain histone lysine methyltransferase that has previously been shown to interact with nuclear receptors. We describe here a novel NSD1-interacting protein, Nizp1, that contains a SCAN box, a KRAB-A domain, and four consensus C2H2-type zinc fingers preceded by a unique finger derivative, referred to herein as the C2HR motif. The C2HR motif functions to mediate protein-protein interaction with the cysteine-rich (C5HCH) domain of NSD1 in a Zn(II)-dependent fashion, and when tethered to RNA polymerase II promoters, represses transcription in an NSD1-dependent manner. Mutations of the cysteine or histidine residues in the C2HR motif abolish the interaction of Nizp1 with NSD1 and compromise the ability of Nizp1 to repress transcription. Interestingly, converting the C2HR motif into a canonical C2H2 zinc finger has a similar effect. Thus, Nizp1 contains a novel type of zinc finger motif that functions as a docking site for NSD1 and is more than just a degenerate evolutionary remnant of a C2H2 motif.

Characterization of the SCAN box encoding RAZ1 gene: analysis of cDNA transcripts, expression, and cellular localization

The SCAN box (SRE-ZBP; CT-fin51; AW-1; Number 18) is a highly-conserved 80-amino-acid domain identified in a subset of C2H2 zinc finger proteins. We and others have recently demonstrated that the SCAN box is a protein association domain that mediates hetero- and homo-protein associations with SCAN box containing proteins. RAZ1 (SCAN-related protein associated with MZF1B) is a novel gene identified in a yeast two hybrid genetic screen for binding to the MZF1B SCAN box. RAZ1 maps to chromosome 20q11 at a region frequently disrupted in various leukemias. We characterized the RAZ1 gene by analysing cDNA transcripts, mRNA expression, and cellular localization of the expressed protein. RAZ1 mRNA expression was detected in various human tissues and cell lines by Northern blot analysis and multiple tissue expression arrays. Highest levels of expression are in prostate, testis, thyroid, liver, and kidney. The RAZ1 gene produces two transcripts with variant 5′-untranslated regions containing identical open reading frames that express a 28 kDa protein in vitro. RAZ1 transcription start sites were mapped by primer extension and confirmed by identification of the RAZ1 promoter in the 5′ flanking genomic DNA. RAZ1 protein fused to the green fluorescent protein (GFP) localizes to the nucleus in a diffuse pattern and the carboxyl terminus containing the SCAN-related domain is sufficient for nuclear localization. These data suggest that RAZ1 is a widely expressed nuclear protein that may function as a key regulator of zinc finger transcription factor function.

A novel human SCAN/(Cys)2(His)2 zinc-finger transcription factor ZNF323 in early human embryonic development

The C 2H 2 zinc-finger motif found in many transcription factors is thought to be important for nucleic acid binding and/or dimerization. Here, we have identified and characterized a novel zinc-finger gene named ZNF323 using degenerate primers from an early human embryo heart cDNA library. The predicted protein contains six different C 2H 2 type zinc fingers and a SCAN box. ZNF323 maps to chromosome 6p22.1–22.3. The expression levels were different during different development stages of human embryo between 15 and 23 weeks. Northern blot analysis shows that a 3.2-kb transcript specific for ZNF323 was expressed at high levels in the lung, liver, and kidney, while weakly expressed in intestine, brain, muscle, cholecyst, heart, and pancreas. In adult tissues, ZNF323 is expressed at high levels in liver and kidney, weakly in lung, pancreas, brain, placenta, muscle, and heart. Taken together, these results indicate that ZNF323 is a member of the zinc-finger transcription factor family and may be involved in the development of multiple embryonic organs.

Identification of SCAN dimerization domains in four gene families

Zinc-finger transcription factors are often accompanied by modular sequence motifs such as the Kruppel-associated box (KRAB) and the SCAN domain. The KRAB domain mediates transcriptional repression while the SCAN domain mediates selective protein dimerization. The hypoalphalipoproteinemia susceptibility gene ZNF202 encodes a SCAN box and a KRAB domain followed by eight Cys 2–His 2 zinc-finger motifs. In order to identify the existence of genes which encode proteins of structural homology to ZNF202, a mouse lambda library was screened with a human ZNF202 cDNA probe. The isolated cDNA clones represented three SCAN-domain-encoding gene families. We purified three novel cDNAs that encode a SCAN-KRAB-(Cys 2–His 2) x domain alignment and one cDNA that encodes a SCAN-(Cys 2–His 2) x domain alignment. In addition, we identified one cDNA sequence with a predicted protein sequence containing a KRAB-SCAN-KRAB-(Cys 2–His 2) x domain alignment. Therefore, when combined with the recently discovered family of isolated SCAN-domain-encoding genes, four SCAN domain gene families can be distinguished. The consensus sequences for the murine SCAN and KRAB domains are highly conserved within the mammalian phylogenetic tree which may be useful in elucidating the biological function of these protein modules and the crucial residues responsible for their binding specificity.

Identification and characterization of SKAT-2, a novel Th2-specific zinc finger gene.

We have identified a novel Kruppel-type zinc finger (ZF) gene, SKAT-2, which is selectively expressed by murine Th2 cells. The protein encoded by this gene has 14 C2H2-type ZF tandemly arrayed at its C terminus and N-terminal SCAN box and KRAB domains. SKAT-2 is tissue restricted in expression at the RNA level, detectable only in brain and at low levels in kidney and spleen and few hematopoietic cell lines. By in situ hybridization, SKAT-2 expression was found to peak in antigen-stimulated CD4(+) T cells after 2-3 days of culture under Th2 but not Th1 biasing conditions. This pattern of expression closely mirrored that of GATA-3 in the same cells. In transient transfection experiments in phorbol 12-myristate 13-acetate/ionomycin-stimulated EL4 cells, SKAT-2 was found to up-regulate the activity of the IL-4 but not the IL-5 promoter, contrasting with the ability of GATA-3 to activate both promoters. This result was confirmed using clones of EL4 cells stably expressing an inducible form of SKAT-2, thus SKAT-2 is a novel Th2-specific gene that may play a role in selective regulation of cytokine genes in T cells.

Human myeloid zinc finger gene MZF produces multiple transcripts and encodes a SCAN box protein

The myeloid zinc finger gene 1 ( MZF1) encodes a C 2H 2 zinc finger transcription factor that regulates granulopoiesis and may have a regulatory role in cellular proliferation and oncogenesis. The MZF1 gene has been previously reported to be 3 kb and without introns. However, at least three transcripts of approximately 3, 7.5, and 9 kb are detected by MZF1-specific probes in northern blot analysis and the identity of the transcripts has not been addressed. We screened a K562 cDNA library and identified novel transcripts, MZF1B and MZF1C. The 2.9 kb MZF1B cDNA encodes a putative 734 aa protein and MZF1C maintains an identical open reading frame with 320 nucleotides deleted in the 5′-untranslated region. The MZF1B/1C protein contains all but the first eight amino acids of MZF1. Thus, MZF protein isoforms share 100 aa, as well as the bipartite 13 zinc finger DNA binding domain. In addition, MZF1B/ 1C encodes a unique 257 aa MZF1B/C amino terminus containing a SCAN box, or leucine-rich domain, which has recently been demonstrated to facilitate protein interactions. Sequence analysis reveals that the MZF gene contains six exons and spans 11 kb and may be the most telomeric gene on chromosome 19q13. Exons 1–6 produce MZF1B/ C cDNA, whereas MZF1 cDNA initiates within intron 5 and continues through exon 6. The 7.5 and 9 kb transcripts are incompletely processed and contain intron sequences. These studies are the first description of the complete human MZF gene and of the composition of the multiple transcripts that are detected by northern blot analysis.

Identification of a Novel SCAN Box-related Protein That Interacts with MZF1B: THE LEUCINE-RICH SCAN BOX MEDIATES HETERO- AND HOMOPROTEIN ASSOCIATIONS

The SCAN box or leucine-rich (LeR) domain is a conserved motif found within a subfamily of C(2)H(2) zinc finger proteins. The function of a SCAN box is unknown, but it is predicted to form alpha-helices that may be involved in protein-protein interactions. Myeloid zinc finger gene-1B (MZF1B) is an alternatively spliced human cDNA isoform of the zinc finger transcription factor, MZF1. MZF1 and MZF1B contain 13 C(2)H(2) zinc finger motifs, but only MZF1B contains an amino-terminal SCAN box. A bone marrow cDNA library was screened for proteins interacting with the MZF1B SCAN box domain and RAZ1 (SCAN-related protein associated with MZF1B) was identified. RAZ1 is a novel cDNA that encodes a SCAN-related domain and arginine-rich region but no zinc finger motifs. Co-immunoprecipitation assays demonstrate that the SCAN box domain of MZF1B is necessary for association with RAZ1. By yeast two-hybrid analysis, the carboxyl terminus of RAZ1 is sufficient for interaction with the MZF1B SCAN box. Furthermore, MZF1B and RAZ1 each self-associate in vitro via a SCAN box-dependent mechanism. These data provide evidence that the SCAN box is a protein interaction domain that mediates both hetero- and homoprotein associations.

Molecular Cloning of Six Novel Krüppel-like Zinc Finger Genes from Hematopoietic Cells and Identification of a Novel Transregulatory Domain KRNB

To clone zinc finger genes expressed in hematopoietic system, we designed primers based on conserved Cys(2)/His(2) zinc finger sequences to amplify corresponding domains from mRNA of normal bone marrow and leukemia cell line NB4. DNA fragments of novel zinc finger genes were chosen and used as probe pool to screen cDNA libraries or subject to rapid amplification of cDNA ends in order to obtain full-length cDNA. Six cDNAs including whole open reading frame of zinc finger proteins, named as ZNF191, ZNF253 (BMZF-1), ZNF255 (BMZF-2), ZNF256 (BMZF-3), ZNF257 (BMZF-4), and ZNF254 (BMZF-5) were obtained. All six belong to the Krüppel-like zinc finger gene family, and typical transcriptional regulatory motifs exist in the N-terminal moiety, such as the SCAN box in ZNF191, and the KRAB domains in ZNF253, ZNF254, ZNF256, and ZNF257. A previously undefined sequence nominated as Krüppel-related novel box, which may represent a new transregulatory motif, was revealed at the N terminus of ZNF255. The transregulatory function of non-zinc finger regions of ZNF191, ZNF253, and ZNF255 were addressed in yeast and mammalian cells. The results indicated that ZNF255 might be a conditional transactivator, whereas ZNF253 and ZNF191 displayed a suppressive effect on the transcription in yeast and/or mammalian systems.

The zinc finger-associated SCAN box is a conserved oligomerization domain.

A number of Cys(2)His(2) zinc finger proteins contain a highly conserved amino-terminal motif termed the SCAN domain. This element is an 80-residue, leucine-rich region that contains three segments strongly predicted to be alpha-helices. In this report, we show that the SCAN motif functions as an oligomerization domain mediating self-association or association with other proteins bearing SCAN domains. These findings suggest that the SCAN domain plays an important role in the assembly and function of this newly defined subclass of transcriptional regulators.

Isolation, Characterization, and Mapping of a Zinc Finger Gene, ZFP95, Containing Both a SCAN Box and an Alternatively Spliced KRAB A Domain

A new zinc finger gene of the Krüppel family was identified by screening a human fetal cartilage cDNA library with degenerate oligonucleotides. Sequence analysis indicates that ZFP95 contains 12 highly conserved zinc finger motifs at the C-terminus and a SCAN box as well as a KRAB A domain at the N-terminus of the protein. ZFP95 represents a member of a new subclass of Krüppel zinc finger proteins containing both a SCAN box and a KRAB domain. Sequence comparison revealed that ZFP95 is the human ortholog of murine Zfp95, which is differentially expressed during spermatogenesis. We demonstrate that ZFP95 is ubiquitously expressed in adult and fetal tissues with the strongest expression in testis. Two transcripts, 4.2 and 4.6 kb, were detected in all tissues tested. In testis, a third transcript of 3.8 kb was present. RT-PCR analysis confirmed alternative splicing for the KRAB A domain and an upstream exon leading to three transcripts of ZFP95 with and without this transcriptional repressor domain. Finally, we show that ZFP95 maps on human chromosome 7q22 between the markers D7S651 and WI-5853.

Three Genes Encoding Zinc Finger Proteins on Human Chromosome 6p21.3: Members of a New Subclass of the Krüppel Gene Family Containing the Conserved SCAN Box Domain

Five genes encoding zinc finger proteins of the Cys2His2(or Krüppel) family were identified by direct cDNA hybridization to YACs 753H12 and 638D7, which encompass a region of human chromosome 6p21.3 extending from just centromeric of the microsatellite marker D6S306 to telomeric of D6S1260. The genes span a distance of approximately 1750 kb. The complete cDNA sequence, genomic structure, and tissue distribution of three of the zinc finger proteins, LD65/ZNF165, ZNF192 (previously called LD5-1), and ZNF193, are described. The three zinc finger proteins do not contain either Krüppel-associated box (KRAB) A or KRAB B domain, present in about one-third of all Krüppel-type zinc finger proteins (E. J. Bellefroidet al.,1991,Proc. Natl. Acad. Sci. USA88: 3608–3612). The three zinc finger proteins do contain the conserved SCAN box domain (A. J. Williamset al.,1995,J. Biol. Chem.270: 22143–22152). SCAN boxes are found in eight other genes in the GenBank database, five of which are also in the Krüppel family of zinc finger proteins lacking KRAB A and B domains and thereby define a new subclass of zinc finger proteins. In addition, three polymorphisms were identified in ZNF192, one of the zinc finger proteins. One of the three polymorphisms, Pro163Leu, is the second proline in a proline cluster (PEPP) in a region separating the SCAN box from the zinc finger motifs.

Isolation and Characterization of a Novel Zinc-finger Protein with Transcriptional Repressor Activity

To identify genes that can repress the expression of growth regulatory molecules, a human fetal cDNA library was screened with a degenerate oligonucleotide that corresponds to the conserved stretch of 6 amino acids connecting successive zinc-finger regions in the Wilms' tumor suppressor/Egr-1 family of DNA-binding proteins. One clone, designated zinc-finger protein 174 (ZNF174), corresponds to a putative transcription factor with three zinc fingers and a novel finger-associated domain, designated the SCAN box. The three Cys2-His2-type zinc fingers are positioned at the carboxyl terminus, while the 65-amino acid finger-associated SCAN box is located near the amino terminus. Chromosomal localization using somatic cell hybrid analysis and fluorescent in situ hybridization mapped the gene for ZNF174 to human chromosome 16p13.3. The 2.5-kilobase transcript from this gene is expressed in a variety of human organs, but most strongly in adult testis and ovary. Fusion of the upstream regulatory region of ZNF174 to the DNA-binding domain of GAL4 revealed that the gene could confer a repression function on the heterologous DNA-binding domain. ZNF174 selectively repressed reporter activity driven by the platelet-derived growth factor-B chain and transforming growth factor-beta 1 promoters and bound to DNA in a specific manner. This member of the C2H2-type zinc-finger family is a novel transcriptional repressor.