Btf and TRAP150 have distinct roles in regulating subcellular mRNA distribution

Area of Science:

• Molecular Biology
• Gene Expression Regulation
• RNA Processing and Export

Background:

• Transcription-coupled pre-mRNA processing, mRNP assembly, and nuclear export are coordinated in mammalian cells.
• Btf (BCLAF1) and TRAP150 (THRAP3) are SR proteins associated with spliceosomes and in vitro spliced mRNPs, suggesting roles in pre-mRNA processing.
• The functional overlap between Btf and TRAP150 remains unclear despite their sequence similarity.

Purpose of the Study:

• To investigate the localization and function of Btf and TRAP150 during gene expression in mammalian cells.
• To determine if Btf and TRAP150 have overlapping or distinct roles in mRNA processing and export.
• To elucidate the specific contribution of Btf to mRNA subcellular distribution.

Main Methods:

• Localization studies of Btf and TRAP150 at reporter gene loci (β-tropomyosin and U2OS 2-6-3) dependent on RNA Polymerase II transcription.
• Analysis of co-localization with reporter RNA, pre-mRNA processing factors, and the exon junction complex (EJC) protein Magoh.
• Assessment of nuclear/cytoplasmic mRNA distribution following Btf or TRAP150 knockdown using quantitative analysis of reporter and endogenous polyadenylated RNAs.

Main Results:

• Both Btf and TRAP150 localize to active reporter gene loci in a transcription-dependent manner.
• Btf and TRAP150 show precise co-localization with the EJC protein Magoh at transcription sites.
• Depletion of Btf, but not TRAP150, leads to increased cytoplasmic accumulation of reporter and endogenous mRNAs, indicating a role in mRNA export.

Conclusions:

• Btf and TRAP150 are recruited to active transcription sites and associate with the exon junction complex.
• Btf plays a critical role in regulating mRNA subcellular distribution, distinct from TRAP150.
• Btf functions in ensuring proper mRNA export from the nucleus in human cells.