Regulation of LIM-domain-binding 1 protein expression by ubiquitination of Lys134.
LDB1 (LIM-domain-binding 1) is a cofactor that participates in formation of transcriptional regulatory complexes involving transcription factors containing LIM domains as well as other factors. The amount of LDB1 protein in cells has previously been shown to be modulated by RNF12 (RING finger protein 12). RNF12 is an E3 ubiquitin ligase that can target LDB1 for poly-ubiquitination and degradation via the proteasome. We find that in HEK (human embryonic kidney)-293 cells expression of RNF12 leads to mono-ubiquitination of LDB1 and increased levels of LDB1 protein. Mutagenesis studies identified Lys134 of LDB1 as the residue that is mono-ubiquitinated by RNF12. Mutation of Lys134 of LDB1 to arginine blocks the formation of mono-ubiquitinated LDB1 and surprisingly also increases LDB1 protein expression in HEK-293 cells. This leads to a model in which Lys134 of LDB1 can be either mono-ubiquitinated, leading to stabilization, or poly-ubiquitinated, leading to degradation by the proteasome pathway. We also find that ubiquitin-LDB1 fusion proteins are stabilized in HEK-293 cells, offering further evidence that mono-ubiquitination stabilizes LDB1 in these cells. Expression in Xenopus laevis embryos of an LDB1 protein in which Lys134 is replaced with arginine leads to enhanced expression of the mutant protein as compared with the wild-type protein. These findings provide evidence that modification of Lys134 can play a major role in regulating LDB1 expression.
Howard
Paul W
PW
Department of Cell and Developmental Biology Oregon Health and Science University, Portland, OR 97239, USA.
Jue
Shall F
SF
Ransom
David G
DG
Maurer
Richard A
RA
D000818:Animals; D002460:Cell Line; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000502:physiology*; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D060588:LIM Domain Proteins; D008239:Lysine / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D011485:Protein Binding / Q000502:physiology; D014157:Transcription Factors / Q000502:physiology*; D054875:Ubiquitination / Q000502:physiology*; D014982:Xenopus laevis
D004268:DNA-Binding Proteins; C486312:LDB1 protein, human; D060588:LIM Domain Proteins; D014157:Transcription Factors; D008239:Lysine