Three Scan Domain Zinc Finger Protein Gene Znf191, Znf396 And Znf397

Many zinc finger genes encoding the SCAN domains are clustered in human genome, e.g. 17p12-13 (ZNF18, ZNF29, ZNF62 and zfp3). One of the SCAN domain genes, ZNF191 has been cloned and mapped to 18q12 . We have identified two novel human SCAN domain genes ZNF396 and ZNF397, which are clustered in the chromosome 18q12: 32850-33000k regions with ZNF191 and ZNF271 genes. It is firstly characterized for the SCAN-(C2H2)X genes to truncate the zinc finger domains at the carboxyl terminus. The unique alternative splicing makes ZNF191, ZNF396 and ZNF397 genes encode two distinct proteins respectively. Three isoforms encoding zinc finger regions are designated as ZNF191-fu, ZNF396-fu and ZNF397-fu (full zinc finger) while the other isoforms truncating zinc finger regions are designated as ZNF191-nf, ZNF396-nf and ZNF397-nf (no zinc finger). Tissue Northern blots (MTN I and MTN II) from 16 adult human tissues are respectively hybridized with isotope-labeled ZNF191-nf, ZNF396-nf or ZNF397-nf cDNA. Three isoforms of ZNF191 transcript, 2.0-kb, 3.7-kb and 7.5-kb, are expressed in testis. Three isoforms of ZNF396 transcript, 1.5-kb, 2.5-kb and 3.9-kb, are highly expressed in liver. Four isoforms of ZNF397 transcript, 1.7-kb, 2.5-kb, 7.0-kb and 9.0-kb, are expressed in a variety of tissues, but expression levels vary dramatically. ZNF191-fu consists of a SCAN domain at N-terminus and four tandem C2H2 zinc finger repeats at C-terminus. ZNF396-fu encoding 335 amino acids consists of a SCAN domain (aa46-141) at N-terminus and three consecutive C2H2 zinc finger repeats at C-terminus (aa251-273, aa279-301, aa307-329). ZNF397-fu encoding 534 amino acids consists of a SCAN domain (aa44-139) at N-terminus and nine consecutive C2H2 zinc finger repeats at C-terminus (aa285-307, aa313-335, aa341-363, aa369-391, aa397-419, aa425-447, aa453-475, aa481-503, aa509-531). ZNF191-nf, ZNF396-nf and ZNF397-nf encode 193, 210 and 198 amino acids with a SCAN domain only respectively. ZNF191-fu, ZNF191-nf, ZNF396-fu, ZNF396-nf, ZNF397-fu or ZNF397-nf can homo-associate, ZNF191-fu can hetero-associates with ZNF191-nf,so does ZNF396-fu with ZNF396-nf, or ZNF397-fu with ZNF397-nf. Moreover, ZNF397-fu can associate with SCAN-domain-containing polypeptide ofZNF397-fu (aa3-250) but not with non-SCAN-domain polypeptide of ZNF397-fu (aa224-534). ZNF191-nf, ZNF396-nf and ZNF397-nf polypeptides diffuse in the whole cell, but ZNF191-fu, ZNF396-fu and ZNF397-fu polypeptides target specifically to the nuclei, consistent with the DAPI-staining nuclei. If ZNF191-fu and ZNF191-nf are co-transfected into the cells, ZNF191-nf tends to localize to the nuclei while ZNF191-fu still targets to the nuclei. In the deletion experiment, the SCAN-domain-containing polypeptide of ZNF397-fu (aa3-250) has the same sub-cellular localization pattern in cells as that of ZNF397-nf, and non-SCAN-domain polypeptide of ZNF397 (aa224-534) targets to the nuclei just like that of ZNF397-fu. It is inferred that the nuclear localization signal of ZNF397-fu might be within zinc finger regions (aa251-534). All six isoforms except for ZNF397-fu are shown to repress the transcription of reporter gene, but ZNF191-fu and ZNF397-nf have the stronger repression activity. Without its nine zinc finger repeats, ZNF397-fu is a transcriptional activation factor. Through the series deletion, the different sub-domains of ZNF191-fu contribute to the distinct effects on the transcription repression. At last, the central fragment of ZNF191-fu (aa139-247) fused with the GST tag is used to immunize rabbit, and the purified polyclonal antibody can detect both transfected and endogeneous ZNF191 proteins.