Mammalian cells express two differently localized Bag-1 isoforms generated by alternative translation initiation

Area of Science:

• Molecular Biology
• Cell Biology
• Biochemistry

Background:

• The Bcl-2 oncoprotein regulates apoptosis, and Bag-1 protein interacts with Bcl-2 to inhibit this process.
• Bag-1 protein is also known as RAP46 and interacts with steroid hormone receptors.
• Confusion exists regarding the structure of Bag-1/RAP46 proteins and their relationship to endogenous Bag-1.

Purpose of the Study:

• To characterize Bag-1 expression in mammalian cells.
• To clarify the structure and origin of different Bag-1 isoforms.
• To investigate the functional implications of distinct Bag-1 isoforms.

Main Methods:

• Characterization of Bag-1 expression in mammalian cells.
• Sequencing of murine Bag-1 RNA 5' sequences.
• Determination of Bag-1 isoform translation initiation sites in human cells.

Main Results:

• Two Bag-1 isoforms, 50 kDa (p50) and 36 kDa (p36), were identified in human cells, in addition to the known 32 kDa murine isoform.
• p50 and p32 Bag-1 proteins exhibit distinct subcellular localizations, suggesting functional differences.
• Human and murine Bag-1 cDNAs contain longer open reading frames than previously thought.
• p50 Bag-1 initiates translation at an upstream CUG codon, while p36 Bag-1 initiates at a downstream AUG codon.
• Cells do not express Bag-1/RAP46 proteins with previously proposed structures.

Conclusions:

• Mammalian cells express two Bag-1 isoforms (p50 and p36) generated by alternative translation initiation.
• These isoforms possess distinct subcellular localizations, implying diverse functional roles.
• Bag-1 proteins may play a dual role in regulating both apoptosis and steroid hormone-dependent transcription.