The transactivation domain of heterogeneous nuclear ribonucleoprotein K overlaps its nuclear shuttling domain.
The heterogeneous nuclear ribonucleoprotein K (hnRNP K) protein is a versatile molecule that interacts with RNA, DNA, and a number of transcription factors, implicating it in transcription, splicing, and translation processes. The underlying mechanism of transcription stimulation by hnRNP K is not well understood. To explore the possibility of a putative transactivation activity of hnRNP K, we produced constructs in which the yeast Gal4 DNA-binding domain was fused to various hnRNP K fragments in one-hybrid mammalian cells. Our results reveal that the K nuclear shuttling (KNS) domain, a well-known signal for nuclear import and export, is also responsible for the transactivation activity of hnRNP K protein. Importin alpha and beta proteins are involved in the regulation of the transactivation activity of the KNS domain via their competition for the nuclear pore complex. Site-directed mutants of serine residue 353 to alanine or aspartic acid or a series of truncated mutants of amino acids 338-363 of hnRNP K suggest the transactivation activity of KNS is primarily dependent on its amino acid composition and intact structure. Our results suggest that endogenous p53 is not required for the activity of the KNS domain, but that overexpression of exogenous p53 might affect its activity in a dose-dependent manner. We thus demonstrate the existence of a strong transactivation domain in hnRNP K and define the regulatory mechanism involved in its protein-protein interaction within the KNS domain in cells.
Chan
James Yi-Hsin
JY
Graduate Institute of Medical Sciences, National Defense Medical Center, 161, Section 6, MinChuan East Road, Taipei 114, Taiwan, ROC. jyhchan@ndmctsgh.edu.tw
Huang
Shih-Ming
SM
Liu
Shu-Ting
ST
Huang
Chun-Hung
CH
D000595:Amino Acid Sequence; D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism*; D006367:HeLa Cells; D034641:Heterogeneous-Nuclear Ribonucleoprotein K / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D049790:Nuclear Export Signals; D010768:Phosphoserine / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D015533:Transcriptional Activation / Q000235:genetics*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D028901:alpha Karyopherins / Q000378:metabolism; D028961:beta Karyopherins / Q000378:metabolism
D034641:Heterogeneous-Nuclear Ribonucleoprotein K; D049790:Nuclear Export Signals; D016159:Tumor Suppressor Protein p53; D028901:alpha Karyopherins; D028961:beta Karyopherins; D010768:Phosphoserine