The transcriptional control of a newly identified potassium channel gene, Kv3.2
The completion of the human genome project revealed the existence of far more non-coding sequence (sequence that does not code for protein) than was originally anticipated. While these regions were once dismissed as “junk DNA,” recent research suggests that a large portion of this material is dedicated to transcriptional control – when, where, and under what conditions a gene is expressed. The goals of this project are twofold: 1) To examine the transcriptional control of a novel potassium channel gene in Drosophila – Kv3.2. 2) To look for any commonality in the regulatory elements (the noncoding sequences that control gene expression) of Kv3.2 and the closest related gene Kv3.1. 5 The first part of this project was determining where in the fruit fly the Kv3.2 ion channel is made. This task required a technique called in situ hybridization. A probe was produced specific to the Kv3.2 gene product and a chemical reaction revealed where in the fly the probe had found its target. It was seen that Kv3.2 is solely expressed in the central nervous system of Drosophila embryos. This is identical to the embryonic expression of Kv3.1, suggesting that there may be common regulatory elements controlling the transcription of the two genes. The second part of this project was examining the DNA sequences responsible for regulating the transcription of Kv3.2. There is currently no method for identifying these sequences directly. To do this, we must rely on the natural process of transcription to produce a gene product and then map this back to the genomic DNA. This technique was used to obtain the full-length transcriptional product of the Kv3.2 gene. The predicted transcription start site (TSS, the location where gene expression begins) was corrected and a new TSS was identified some 19,000 bases from the remainder of the gene. Knowing the location of the TSSs allows for more accurate targeting of the sequences that regulate transcription initiation. Sequence analysis was used to search for evolutionarily conserved sequences near the TSSs as potential candidates for regulatory elements. In the post-genomic era, science faces new challenges. Whereas much of the previous effort was in locating genes themselves, the focus is now turning to identifying the regulatory elements between genes. Some of these elements for the related gene, Kv3.1, have already been identified. Similarly, future research will rely on the findings reported here to study the sequences regulating Kv3.2 expression.