Handout 8—Pre-mRNA Gene Therapy for Tazswana

—What is “alternative splicing” and how does it apply to Tazswana’s case?
—Explain how hybridization (complementary base-pairing) of the DNA oligonucleotides to the cryptic and aberrant pre-mRNA sequences would correct the splicing defect.
—If Tazswana’s 5′ splice site at the correct location (exon2/intron2 boundary) had a point mutation from G to U so that the original 5′splice site read “UU” instead of “GU,” would the gene therapy correct her β-thalassemia? Explain your answer.
—(Optional) Since the oligonucleotides target splice site consensus sequences that are common to all introns, wouldn’t they potentially cause side-effects in many cell types? Discuss within your group and come up with:
—If you were in Tazswana’s family, would you want Tazswana to undergo gene therapy treatment? Explain why or why not.

Answers:


1. Alternative splicing is the RNA splicing variation mechanism in which the exons of the pre-mRNA are separated and reconnected so as to produce alternative ribonucleotide arrangements. These linear combinations then undergo the process of translation where specific and unique sequences of amino acids are specified, resulting in isoform proteins. Alternative splicing facilitates the synthesis of a greater variety of proteins.

2. The hybridization of the DNA oligonucleotides would correct the splicing defect since the attach both exons once again, as if the process never happened. When these oligonucleotides do this, they rewrite the nuclear information so the gene splicing would be executed correctly in the correct spots, specifically the 5’ splice site.

3. The gene therapy would correct Miss X’s beta thalassemia if the 5’ splice site is spliced in the correct location, having a point mutation from G to U so that it can read “UU” instead of “GU”. The gene therapy would basically rewrite the sequence, performing the splicing correctly and eliminating the beta thalassemia gene defect from her genetic code.

4. The oligonucleotides that target splice site consensus sequences will probably not cause any side effects. This process has only been done on a small scale and it proved to have no adverse effects on the subject.

5. If we were in Miss X’s family, we would certainly want her to undergo the gene therapy treatment since it would cure a disease that causes many side effects and a poor quality of life. The gene therapy would cure her from the genetic defect, thus helping her lead a normal life.