The Molecular Mechanism Of Mouse HNF1? Regulating Glucose Reabsorption In Renal Epithelial Cells

The kidney is an important organ of the human body,it can maintain electrolyte and acid-base balance.It also has endocrine function.The regulation of glucose homeostasis by the kidney is mainly through gluconeogenesis and glucose reabsorption.Hepatocyte Nuclear Factor 1?(HNF1?)is a transcription factor that can be expressed in the liver,kidney,pancreas,and digestive tract.It plays an important role in the differentiation of kidney and renal tubular epithelial cells,and loss of HNF1? function can also trigger renal glucose metabolism-related diseases such as renal Fanconi Syndrome(FS).HNF1? as a transcription factor requires acetyltransferase as a co-activator to make it function.This subject is based on the biological characteristics of HNF1? to study its molecular mechanism of renal glucose reabsorption.We constructed a mouse HNF1?expression vector,transfected the recombinant plasmid into human embryonic kidney cell line HEK293 T cells and human renal tubular epithelial cell line HK2 cells,and detected the absorption of fluorescent glucose analog 2-NBDG by cells using flow cytometry Situation,it was found that overexpression of HNF1? can promote the absorption of 2-NBDG by cells.Real-time quantitative PCR was used to detect the m RNA levels of HNF1? downstream target genes such as GLUT2,SGLT2,and INSIG1.It was found that overexpression of HNF1? increased the expression levels of kidney-related sugar transporter genes such as GLUT2 and SGLT2.GLUT2 is a glucose transporter.The luciferase reporter gene test found that overexpression of HNF1? enhanced the activity of the GLUT2 reporter gene,and interference with HNF1? inhibited the activity of the GLUT2 reporter gene.Through EMSA and Ch IP experiments,it was found that HNF1? directly binds to the promoter of GLUT2.Early laboratory research found that Lys117,the acetylation site of HNF1?,plays an important role in kidney glucose reabsorption in mice.Using reporter gene experiments,we found that the transcriptional activation ability of the GLUT2 reporter gene was significantly reduced after HNF1?K117 mutation.The histone acetyltransferase PCAF can up-regulate the transcriptional activity of HNF1? in a dose-dependent manner,but the regulatory effect on the K117 mutant is significantly weakened;The results of co-immunoprecipitation experiments showed that the K117 mutation did not affect the protein interaction between PCAF and HNF1?,but the K117 mutation affected the interaction between HNF1? and another acetyltransferase CBP,and PCAF can significantly promote the interaction between HNF1? and CBP.When K117 is mutated,the promotion effect of PCAF is obviously weakened.In summary,HNF1? promotes glucose reabsorption of renal tubular epithelial cells by regulating the transcription activity of GLUT2,which is closely related to the acetylation site K117 of HNF1?.PCAF regulates the transcriptional activity of HNF1? by promoting the protein interaction between HNF1?K117 and CBP.The synergistic regulation of CBP and PCAF on HNF1?K117 provides new ideas for the study of HNF1? function and also provides a new direction for the study of renal glucose metabolism.