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Isoform specific knockdown of the ETS transcription factor Pointed in Drosophila S2 cells.

ABSTRACT: Alternate splicing of the pointed ( pnt ) gene locus produces two major isoforms, PntP1 and PntP2. Understanding their individual contributions to key developmental processes and identification of their genome-wide transcriptional targets has been hampered by a number of factors including their essential roles during embryonic development, and co-expression in several tissues. siRNAs were designed to target isoform-specific exons that code for the unique N-terminal region of either PntP1 or PntP2. The efficacy and specificity of the siRNAs were examined by co-transfection of isoform specific siRNAs with plasmids encoding epitope tagged PntP1 or PntP2 in Drosophila S2 cells. All P1-specific siRNAs were demonstrated to knockdown PntP1 protein level to greater than 95%, while having nominal impact on PntP2 level. Similarly, PntP2 siRNAs while ineffective at eliminating PntP1, were shown to reduce PntP2 protein level by 87-99%.

SUBMITTER: Vivekanand P

PROVIDER: S-EPMC10245148 | biostudies-literature | 2023

REPOSITORIES: biostudies-literature

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Isoform specific knockdown of the ETS transcription factor Pointed in Drosophila S2 cells.

Vivekanand Pavithra P

microPublication biology 20230523

Alternate splicing of the pointed ( pnt ) gene locus produces two major isoforms, PntP1 and PntP2. Understanding their individual contributions to key developmental processes and identification of their genome-wide transcriptional targets has been hampered by a number of factors including their essential roles during embryonic development, and co-expression in several tissues. siRNAs were designed to target isoform-specific exons that code for the unique N-terminal region of either ...[more]

PMID: 37292519

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