Related Experiment Videos
Exon skipping by mutation of an authentic splice site of c-kit gene in W/W mouse
S Hayashi1, T Kunisada, M Ogawa
1Department of Pathology, Kumamoto University Medical School, Japan.
Abstract:
The murine mutation dominant white spotting (W) is in the proto-oncogene, c-kit. The receptor tyrosine kinase encoded by this gene has pleiotropic effects on murine development including hemopoietic cells, pigment cells, and germ cells. In this study, mutation in W homozygous mouse was identified as a single base substitution (GT----AT) at the 5'-splice donor site of the exon which encodes the transmembrane domain. Two types of aberrant exon skipping resulted from this mutation, occurred in a tissue specific manner. Either transcript lost the exon coding for transmembrane region and therefore the product might not be functional for signal transduction. Any unusual cryptic splice sites were not activated by this mutation as beta-globin gene in beta-thalassaemia. In addition, twelve base pair sequence of the 3'-end of the exon prior to the exon coding for transmembrane domain was found to be alternatively spliced. These findings should provide the genetic base for not only the receptor function but the splicing mechanism.
Insights
The white spotting (W) mutation in mice, affecting the c-kit proto-oncogene, disrupts transmembrane signaling. This genetic study identifies a specific splice site mutation causing aberrant exon skipping, impacting receptor tyrosine kinase function.
Area of Science:
- Molecular Biology
- Genetics
- Developmental Biology
Background:
- The murine dominant white spotting (W) mutation is linked to the c-kit proto-oncogene.
- The c-kit receptor tyrosine kinase plays crucial roles in murine development, affecting hematopoietic, pigment, and germ cells.
Purpose of the Study:
- To identify the specific genetic mutation responsible for the W mutation in homozygous mice.
- To investigate the molecular mechanisms underlying the aberrant splicing caused by this mutation.
- To understand the impact of the mutation on c-kit receptor function and signaling.
Main Methods:
- Genetic sequencing to identify the base substitution in the W mutation.
- Analysis of mRNA transcripts to detect aberrant splicing patterns.
- Comparison of splicing defects with known mutations, such as in beta-thalassemia.
Main Results:
- A single base substitution (GT----AT) was identified at the 5'-splice donor site of the exon encoding the transmembrane domain.
- Two types of tissue-specific aberrant exon skipping were observed.
- The mutation resulted in the loss of the exon coding for the transmembrane region, potentially impairing signal transduction.
- Alternative splicing of a 12-bp sequence at the 3'-end of the preceding exon was also detected.
- No activation of cryptic splice sites was observed.
Conclusions:
- The identified splice site mutation provides a genetic basis for the W mutation's effects.
- The findings elucidate the role of the transmembrane domain in c-kit receptor function.
- This study sheds light on the complex mechanisms of alternative splicing in mammalian development.