Mammalian cells have two functional RCC1 proteins produced by alternative splicing

Area of Science:

• Molecular Biology
• Genetics
• Cell Biology

Background:

• Previous cloning of human RCC1 cDNAs identified variations in noncoding regions.
• RCC1 (Regulator of Chromosome Condensation 1) is crucial for cell division and gene regulation.

Purpose of the Study:

• To identify and characterize novel human and hamster RCC1 cDNAs.
• To investigate the functional significance of newly discovered RCC1 coding regions.
• To explore the evolutionary conservation of RCC1 gene regulation.

Main Methods:

• Cloning of new human and hamster RCC1 cDNAs.
• Complementation assays using the tsBN2 cell line to test RCC1 function.
• Sequence analysis to identify insertions and compare coding regions.
• Genomic sequence analysis to locate inserted sequences and identify splice sites.

Main Results:

• New human and hamster RCC1 cDNAs were identified, encoding a protein variant (RCC1-I) with an amino acid insertion in the N-terminal region.
• The novel RCC1-I variant could complement the RCC1 mutation in tsBN2 cells, indicating functional relevance.
• The insertion site was mapped to a novel exon (6') located between the 6th and 7th exons of the human RCC1 gene.
• Evidence of alternative splicing producing RCC1-I mRNA was found, with conserved splice sites (GT-AG) in the 6' exon.
• The presence of the insertion at the same site in both human and hamster RCC1 genes suggests evolutionary conservation.

Conclusions:

• Alternative splicing of the RCC1 gene generates functional protein variants (RCC1-I).
• The mechanism of alternative splicing in the RCC1 gene is conserved between humans and hamsters.
• This study expands the understanding of RCC1 diversity and its evolutionary regulation.