Alternative translation initiation produces synaptic organizer proteoforms with distinct localization and functions

Area of Science:

• Molecular Biology
• Neuroscience
• Proteomics

Background:

• Many messenger RNAs (mRNAs) possess multiple translation initiation sites (TISs), leading to diverse protein isoforms (proteoforms).
• The functional significance of alternative TISs and their resulting proteoforms is largely unknown, particularly in neuronal contexts.

Purpose of the Study:

• To investigate the functional consequences of alternative TIS usage in neuronal pentraxin receptor (NPR) mRNA.
• To explore the role of RNA secondary structure and neuronal activity in regulating proteoform ratios.
• To determine the impact of altered proteoform ratios on synaptic function and behavior.

Main Methods:

• Analysis of alternative CUG and AUG translation initiation sites in NPR mRNA.
• Investigation of RNA secondary structure and neuronal activity's influence on proteoform ratios.
• Functional assays involving AMPA-type glutamate receptor clustering and behavioral studies in mice.

Main Results:

• Alternative TIS usage in NPR mRNA generates two proteoforms, with ratios modulated by RNA structure and neuronal activity.
• A secreted NPR proteoform, produced by downstream AUG initiation, promotes synaptic clustering of AMPA receptors.
• Altered NPR proteoform ratios impair AMPA receptor levels in interneurons and affect learning behaviors.

Conclusions:

• Alternative TIS usage provides a mechanism for generating functional protein diversity, impacting synaptic plasticity and behavior.
• The study identifies plasticity of N-terminal signal sequences, regulated by alternative TISs, as a key factor in protein localization and function diversification.
• Findings highlight a potentially widespread mechanism for regulating protein function through alternative mRNA translation.