Transcriptional Control of the Human Sodium-coupled Neutral Amino Acid Transporter System A Gene by Amino Acid Availability Is Mediated by an Intronic Element

Abstract

System A amino acid transporter (SNAT2) gene expression is up-regulated at the transcriptional level in response to amino acid deprivation. Functional analysis of genomic fragments 5′ upstream of the transcription start site, for both human and mouse SNAT2 genes showed that these regions exhibit promoter activity, but were amino acid unresponsive. However, when the human and mouse constructs were extended to include intron 1, it was observed that the rate of transcription was increased following amino acid deprivation. Deletion analysis of the human gene identified an intron 1 that was for amino to SNAT2 of the the and it was that of these an amino acid response to of is to the in the human SNAT2 in that it in an and and it transcription to System A amino acid transporter (SNAT2) gene expression is up-regulated at the transcriptional level in response to amino acid deprivation. Functional analysis of genomic fragments 5′ upstream of the transcription start site, for both human and mouse SNAT2 genes showed that these regions exhibit promoter activity, but were amino acid unresponsive. However, when the human and mouse constructs were extended to include intron 1, it was observed that the rate of transcription was increased following amino acid deprivation. Deletion analysis of the human gene identified an intron 1 that was for amino to SNAT2 of the the and it was that of these an amino acid response to of is to the in the human SNAT2 in that it in an and and it transcription to System A amino acid is of amino and for amino of that the of amino acid and transporter System A the for and to of is to the and rate the that the exhibit A to System A is up-regulated in in response to amino acid but was the of the of that the gene System A is at the level of transcription the for System and the and amino acid it was that human SNAT2 is the amino acid and is SNAT2 expression and System A amino transcription human SNAT2 genomic A in amino acid in of and transcription of A of of increased transcription for genes and is the human gene for transcription in response to amino acid that response and and response amino acid acid response response amino acid acid response in the both of these human promoter that the and an amino acid response site, it both of these of the of transcription for increased SNAT2 following amino acid that for and were observed the for when were in amino was of an in was the of SNAT2 was following amino acid was for the the in SNAT2 was of these it was that it is that the SNAT2 gene the genomic the of is to the of the SNAT2 and the intron for both the human and the mouse to the that to SNAT2 is that to the transcription and the is that the of the that the activity, transcription to promoter and in and were in at in of was and amino acid the was and were to in to the amino acid in the and was the was to analysis for the human SNAT2 and the were were were and of was and to the of the for was in expression the of and SNAT2 genomic to the 5′ upstream of the transcription start site, were the were in of the the of the A mouse genomic the was mouse genomic the human SNAT2 the at and the at of the human gene of the mouse genomic of was for the and to the mouse were and showed that the mouse transcription start it was that the of the mouse fragments were and promoter was that were the the to the transcriptional of these genomic were 5′ upstream promoter of the human SNAT2 gene to of the and were and and in were for mouse and human SNAT2 genomic fragments and the were and was to the of the and to the human in and were in at the of and for was 1 of and the was at of of of to the human the promoter in of the was for the amino acid response of to for were and the was the were and for in 1 of the of the were of and to to of the and were in the System the the human SNAT2 was was the and of the constructs was in in and for were for of and were of the were and for in both was was were of of 1 in of was to for and to to in were of in the of of was of of and for was at of and were at the was at for were to analysis and of was to 5′ of were of in the of the was the the in and of the SNAT2 gene were the to genomic for SNAT2 and and and and genomic for the and human SNAT2 genes were the to the and in to genomic was to the SNAT2 gene for and the gene of and 1 of the start and in and following were for analysis of SNAT2 was in human and SNAT2 was increased in of to the the System A and amino acid of transcription that in these were the for the in human were for in amino analysis and was and were for SNAT2 of the analysis was of the and and the SNAT2 in to the genomic for the amino of the human SNAT2 fragments of promoter were the that the of transcription in amino promoter was in the human SNAT2 upstream genomic fragments in response to amino acid the promoter in the amino transcription was observed when human SNAT2 genomic was that the amino acid response of the human SNAT2 gene in the 5′ upstream of mouse genomic in amino transcription the SNAT2 5′ upstream in response to amino acid deprivation. were constructs the genomic fragments of the human mouse SNAT2 5′ upstream human promoter to was the were for in and the were and an SNAT2 in the the of an of the transcription start site, constructs were the mouse gene that promoter genomic were the and to transcription was A the the 1, and of intron 1 but in transcription following deprivation. Deletion in the 5′ constructs that but rate of transcription following amino acid to in of the the regions of the human gene were were human promoter was to of of the in of the upstream human SNAT2 promoter in amino Deletion of the to in of the amino acid the that both the human and mouse SNAT2 genes an that is in intron 1 for the human the acid in the intron of the SNAT2 were constructs the genomic fragments of the mouse human SNAT2 the were for in and the were and SNAT2 for that the of the human SNAT2 gene was the regions the and were regions were 1 and the that is to the human and the and is of that to the genomic for the mouse and in these for amino of these were in the of the human SNAT2 genomic an of in response to of the but the rate to of the the the amino acid for to for of the human However, when the was was observed in the of the transcription was the the was for the amino it the of transcription when the were the the was for gene expression of SNAT2 gene of the SNAT2 intron 1 and was and regions and to the to the human gene of the regions to the transcription start and the of 1 and intron 1 in of the and the human SNAT2 intron were in the of the SNAT2 genomic to the and to and the the of the to the transcription start of the human were the constructs and the were for in and the were of the and constructs in of the for the was to 1, and both and were the SNAT2 the following fragments were following the human SNAT2 promoter in the expression the the and the and was and the was the human SNAT2 promoter to transcription the was increased the and the of the of the the the expression of the However, to the the of the in the in of transcription following the but to that for the the the of and that the the is to of and in rate of SNAT2 expression when analysis of the human SNAT2 genomic SNAT2 to were the gene the SNAT2 promoter to and the following fragments intron was the were for in and were the of for the was to 1, and were the of the transcription in amino acid the for of the constructs was to 1 to the following SNAT2 an the were in the SNAT2 promoter in to the transcription start site, but 5′ to the in the and to the in both and the of the to the transcription start site, but 5′ to the in the the of in response to amino acid However, of the amino of the of the rate and the were the of the in the in amino acid of was the the SNAT2 were in upstream of the promoter of the in both and the of the the promoter amino acid that the SNAT2 human SNAT2 were the gene to the SNAT2 the were for in and the were the in the SNAT2 was in the SNAT2 promoter in both and and and in the site, is to the promoter and 5′ to the and the site, is to the SNAT2 promoter was the of the SNAT2 was in the gene the the was in both and and and in the site, is 5′ to the and the site, is to the was the in response to was to for is the of the human SNAT2 in the of of these an of but an in were the and were increased in the of was in 1 and and were the SNAT2 was that the in were the analysis of the that these for and the for the of the SNAT2 in an of the of the when an was was observed and analysis of the human SNAT2 were in for and of the to the SNAT2 to and is in and the of the of of for and the of of the the and the of of an of the an the for the is in of genes identified for in response to amino acid in and was in that System amino acid was increased when were in amino acid the to the of response following transporter and of to the of the the of genes System A activity, it was that the for the SNAT2 gene was increased following amino acid and that that for System A transporter the the of SNAT2 transcription the following that the in SNAT2 in is of transcriptional the SNAT2 and intron 1 of the human and mouse and analysis it is that the in the SNAT2 gene is the of 5′ upstream of that the that the SNAT2 in that it amino acid to is in the and is that in the SNAT2 and that the of these is increased when amino analysis that the SNAT2 is of the amino of the SNAT2 gene is the of the genes the amino acid human and human in the promoter the response to amino acid the of and that the of these the the gene an that the of the the that the to to the response the promoter to the the of the SNAT2 the the in the SNAT2 gene is the of the is upstream of the that the SNAT2 is in the but the of upstream of the at the and that upstream of the SNAT2 these in the amino acid of the for the SNAT2 to the of System A transporter the genomic for amino acid of SNAT2 A of the that expression the SNAT2 gene and of the transcriptional that these of System A in to System A expression is an to the of of and to the of in System A amino acid is of amino and for amino of that the of amino acid and transporter System A the for and to of is to the and rate the that the exhibit A to System A is up-regulated in in response to amino acid but was the of the of that the gene System A is at the level of transcription the for System and the and amino acid it was that human SNAT2 is the amino acid and is SNAT2 expression and System A amino transcription human SNAT2 genomic A in amino acid in of and transcription of A of of increased transcription for genes and is the human gene for transcription in response to amino acid that response and and response amino acid acid response response amino acid acid response in the both of these human promoter that the and an amino acid response site, it both of these of the of transcription response amino acid amino acid response response amino acid amino acid response for increased SNAT2 following amino acid that for and were observed the for when were in amino was of an in was the of SNAT2 was following amino acid was for the the in SNAT2 was of these it was that it is that the SNAT2 gene the genomic the of is to the of the SNAT2 and the intron for both the human and the mouse to the that to SNAT2 is that to the transcription and the is that the of the that the activity, transcription to promoter and in and were in at in of was and amino acid the was and were to in to the amino acid in the and was the was to analysis for the human SNAT2 and the were were were and of was and to the of the for was in expression the of and SNAT2 genomic to the 5′ upstream of the transcription start site, were the were in of the the of the A mouse genomic the was mouse genomic the human SNAT2 the at and the at of the human gene of the mouse genomic of was for the and to the mouse were and showed that the mouse transcription start it was that the of the mouse fragments were and promoter was that were the the to the transcriptional of these genomic were 5′ upstream promoter of the human SNAT2 gene to of the and were and and in were for mouse and human SNAT2 genomic fragments and the were and was to the of the and to the human in and were in at the of and for was 1 of and the was at of of of to the human the promoter in of the was for the amino acid response of to for were and the was the were and for in 1 of the of the were of and to to of the and were in the System the the human SNAT2 was was the and of the constructs was in in and for were for of and were of the were and for in both was was were of of 1 in of was to for and to to in were of in the of of was of of and for was at of and were at the was at for were to analysis and of was to 5′ of were of in the of the was the the in and of the SNAT2 gene were the to genomic for SNAT2 and and and and genomic for the and human SNAT2 genes were the to the and were in at in of was and amino acid the was and were to in to the amino acid in the and was the was to analysis for the human SNAT2 and the were were were and of was and to the of the for was in expression the of and SNAT2 genomic to the 5′ upstream of the transcription start site, were the were in of the the of the A mouse genomic the was mouse genomic the human SNAT2 the at and the at of the human gene of the mouse genomic of was for the and to the mouse were and showed that the mouse transcription start it was that the of the mouse fragments were and promoter was that were the the to the transcriptional of these genomic were 5′ upstream promoter of the human SNAT2 gene to of the and were and and and were in at the of and for was 1 of and the was at of of of to the human the promoter in of the was for the amino acid response of to for were and the was the were and for in 1 of the of the were of and to to of the and were in the System the the human SNAT2 was was the and of the constructs was in in and for were for of and were of the were and for in both was was were of of 1 in of was to for and to to in were of in the of of was of of and for was at of and were at the was at for were to and of the SNAT2 gene were the to genomic for SNAT2 and and and and genomic for the and human SNAT2 genes were the to the and in to genomic was to the SNAT2 gene for and the gene of and 1 of the start and in and following were for analysis of SNAT2 was in human and SNAT2 was increased in of to the the System A and amino acid of transcription that in these were the for the the SNAT2 in to the genomic for the amino of the human SNAT2 fragments of promoter were the that the of transcription in amino promoter was in the human SNAT2 upstream genomic fragments in response to amino acid the promoter in the amino transcription was observed when human SNAT2 genomic was that the amino acid response of the human SNAT2 gene in the 5′ upstream of mouse genomic in amino transcription the SNAT2 5′ upstream in response to amino acid deprivation. were constructs the genomic fragments of the human mouse SNAT2 5′ upstream human promoter to was the were for in and the were and an SNAT2 in the the of an of the transcription start site, constructs were the mouse gene that promoter genomic were the and to transcription was A the the 1, and of intron 1 but in transcription following deprivation. Deletion in the 5′ constructs that but rate of transcription following amino acid to in of the the regions of the human gene were were human promoter was to of of the in of the upstream human SNAT2 promoter in amino Deletion of the to in of the amino acid the that both the human and mouse SNAT2 genes an that is in intron 1 for the human the acid in the intron of the SNAT2 were constructs the genomic fragments of the mouse human SNAT2 the were for in and the were and SNAT2 for that the of the human SNAT2 gene was the regions the and were regions were 1 and the that is to the human and the and is of that to the genomic for the mouse and in these for amino of these were in the of the human SNAT2 genomic an of in response to of the but the rate to of the the the amino acid for to for of the human However, when the was was observed in the of the transcription was the the was for the amino it the of transcription when the were the the was for gene expression of SNAT2 gene of the SNAT2 intron 1 and was and regions and to the to the human gene of the regions to the transcription start and the of 1 and intron 1 in of the and the human SNAT2 intron were in the of the SNAT2 genomic to the and to and the the of the to the transcription start of the human were the constructs and the were for in and the were of the and constructs in of the for the was to 1, and both and were the SNAT2 the following fragments were following the human SNAT2 promoter in the expression the the and the and was and the was the human SNAT2 promoter to transcription the was increased the and the of the of the the the expression of the However, to the the of the in the in of transcription following the but to that for the the the of and that the the is to of and in rate of SNAT2 expression when analysis of the human SNAT2 genomic SNAT2 to were the gene the SNAT2 promoter to and the following fragments intron was the were for in and were the of for the was to 1, and were the of the transcription in amino acid the for of the constructs was to 1 to the following SNAT2 an the were in the SNAT2 promoter in to the transcription start site, but 5′ to the in the and to the in both and the of the to the transcription start site, but 5′ to the in the the of in response to amino acid However, of the amino of the of the rate and the were the of the in the in amino acid of was the the SNAT2 were in upstream of the promoter of the in both and the of the the promoter amino acid that the SNAT2 human SNAT2 were the gene to the SNAT2 the were for in and the were the in the SNAT2 was in the SNAT2 promoter in both and and and in the site, is to the promoter and 5′ to the and the site, is to the SNAT2 promoter was the of the SNAT2 was in the gene the the was in both and and and in the site, is 5′ to the and the site, is to the was the in response to was to for is the of the human SNAT2 in the of of these an of but an in were the and were increased in the of was in 1 and and were the SNAT2 was that the in were the analysis of the that these for and the for the of the SNAT2 in an of the of the when an was was observed and analysis of the human SNAT2 were in for and of the to the SNAT2 to and is in and the of the of of for and the of of the the and the of of an of the an the for the is in SNAT2 in to genomic was to the SNAT2 gene for and the gene of and 1 of the start and in and SNAT2 following were for analysis of SNAT2 was in human and SNAT2 was increased in of to the the System A and amino acid of transcription that in these were the for the the SNAT2 in to the genomic for the amino of the human SNAT2 fragments of promoter were the that the of transcription in amino promoter was in the human SNAT2 upstream genomic fragments in response to amino acid the promoter in the amino transcription was observed when human SNAT2 genomic was that the amino acid response of the human SNAT2 gene in the 5′ upstream of mouse genomic in amino transcription A SNAT2 in the the of an of the transcription start site, constructs were the mouse gene that promoter genomic were the and to transcription was A the the 1, and of intron 1 but in transcription following deprivation. Deletion in the 5′ constructs that but rate of transcription following amino acid to in of the the regions of the human gene were were human promoter was to of of the in of the upstream human SNAT2 promoter in amino Deletion of the to in of the amino acid the that both the human and mouse SNAT2 genes an that is in intron 1 for the human the A SNAT2 for that the of the human SNAT2 gene was the regions the and were regions were 1 and the that is to the human and the and is of that to the genomic for the mouse and in these for amino of these were in the of the human SNAT2 genomic an of in response to of the but the rate to of the the the amino acid for to for of the human However, when the was was observed in the of the transcription was the the was for the amino it the of transcription when the were the the was for gene expression the SNAT2 the following fragments were following the human SNAT2 promoter in the expression the the and the and was and the was the human SNAT2 promoter to transcription the was increased the and the of the of the the the expression of the However, to the the of the in the in of transcription following the but to that for the the the of and that the the is to of and in rate of SNAT2 expression when SNAT2 an the were in the SNAT2 promoter in to the transcription start site, but 5′ to the in the and to the in both and the of the to the transcription start site, but 5′ to the in the the of in response to amino acid However, of the amino of the of the rate and the were the of the in the in amino acid of was the the SNAT2 were in upstream of the promoter of the in both and the of the the promoter amino acid that the SNAT2 the of the human SNAT2 in the of of these an of but an in were the and were increased in the of was in 1 and and were the SNAT2 was that the in were the analysis of the that these for and the for the of the SNAT2 in an of the of the when an was was observed and of genes identified for in response to amino acid in and was in that System amino acid was increased when were in amino acid the to the of response following transporter and of to the of the the of genes System A activity, it was that the for the SNAT2 gene was increased following amino acid and that that for System A transporter the the of SNAT2 transcription the following that the in SNAT2 in is of transcriptional the SNAT2 and intron 1 of the human and mouse and analysis it is that the in the SNAT2 gene is the of 5′ upstream of that the that the SNAT2 in that it amino acid to is in the and is that in the SNAT2 and that the of these is increased when amino analysis that the SNAT2 is of the amino of the SNAT2 gene is the of the genes the amino acid human and human in the promoter the response to amino acid the of and that the of these the the gene an that the of the the that the to to the response the promoter to the the of the SNAT2 the the in the SNAT2 gene is the of the is upstream of the that the SNAT2 is in the but the of upstream of the at the and that upstream of the SNAT2 these in the amino acid of the for the SNAT2 to the of System A transporter the genomic for amino acid of SNAT2 A of the that expression the SNAT2 gene and of the transcriptional that these of System A in to System A expression is an to the of of and to the of in A of genes identified for in response to amino acid in and was in that System amino acid was increased when were in amino acid the to the of response following transporter and of to the of the the of genes System A activity, it was that the for the SNAT2 gene was increased following amino acid and that that for System A transporter the the of SNAT2 transcription the following that the in SNAT2 in is of transcriptional the SNAT2 and intron 1 of the human and mouse and analysis it is that the in the SNAT2 gene is the of 5′ upstream of that the that the SNAT2 in that it amino acid to is in the and is that in the SNAT2 and that the of these is increased when amino analysis that the SNAT2 is A of the amino of the SNAT2 gene is the of the genes the amino acid human and human in the promoter the response to amino acid the of and that the of these the the gene an that the of the the that the to to the response the promoter to the the of the SNAT2 the the in the SNAT2 gene is the of the is upstream of the that the SNAT2 is in the but the of upstream of the at the and that upstream of the SNAT2 these in the amino acid of the for the SNAT2 to the of System A transporter the genomic for amino acid of SNAT2 A of the that expression the SNAT2 gene and of the transcriptional that these of System A in to System A expression is an to the of of and to the of in for the mouse genomic and for of the for and