Structure-Function Analyses of Single Cells by Combining Patch-Clamp Techniques with Reverse Transcription-Polymerase Chain Reaction

Seifert, Gerald; Steinhäuser, Christian

doi:10.1007/978-1-59745-492-6_13

Cited by 1 publication

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“…Normalization was performed using the housekeeping gene PBGD as a reference against the expression of CPEB1 gene transcripts. The transcript level of CPEB1 and the reference gene was calculated using the following equation [43]: X CPEB1 /X PBGD = 2 CT PBGD – CT CPEB1 , where X are the respective input copy numbers and C T the threshold cycle numbers for CPEB1 and PBGD.…”

Section: Methodsmentioning

confidence: 99%

Altered splicing leads to reduced activation of CPEB3 in high-grade gliomas

et al. 2016

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Cytoplasmic polyadenylation element binding proteins (CPEBs) are auxiliary translational factors that associate with consensus sequences present in 3′UTRs of mRNAs, thereby activating or repressing their translation. Knowing that CPEBs are players in cell cycle regulation and cellular senescence prompted us to investigate their contribution to the molecular pathology of gliomas–most frequent of intracranial tumors found in humans. To this end, we performed methylation analyses in the promoter regions of CPEB1-4 and identified the CPEB1 gene to be hypermethylated in tumor samples. Decreased expression of CPEB1 protein in gliomas correlated with the rising grade of tumor malignancy. Abundant expression of CPEBs2-4 was observed in several glioma specimens. Interestingly, expression of CPEB3 positively correlated with tumor progression and malignancy but negatively correlated with protein phosphorylation in the alternatively spliced region. Our data suggest that loss of CPEB3 activity in high-grade gliomas is caused by expression of alternatively spliced variants lacking the B-region that overlaps with the kinase recognition site. We conclude that deregulation of CPEB proteins may be a frequent phenomenon in gliomas and occurs on the level of transcription involving epigenetic mechanism as well as on the level of mRNA splicing, which generates isoforms with compromised biological properties.

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Section: Methodsmentioning

confidence: 99%