Abstract
<jats:p>Neuronal outgrowth to support cognitive functions relies on a timely supply of proteins to distal sites in the neuron. To meet this demand, neurons transport mRNAs and ribosomes in large assemblies called 'neuronal RNA granules'. Ribosomes that integrate into these granules initiate protein synthesis in the soma, stall the process, and then become packaged into clusters that are transported to distal sites, where protein synthesis can be reactivated. The exact mechanisms and structure of these ribosome clusters remain unknown. We combined cryo-electron microscopy and cryo-electron tomography to examine these dense ribosome clusters purified from rat brains, which are included in neuronal RNA granules. We found that these clusters contain ribosomes stalled at various stages of elongation and that the mRNA sequences where the ribosomes stall facilitate stalling. The ribosome clusters contain multiple polysomal units, and ribosomes from nearby polysomes interact through rRNA expansion segments, only seen before in disomes of inactive or hibernating ribosomes in neurons. The ribosomes in the clusters form unique three-dimensional arrays that evade detection by the ribosome-associated quality-control and NO-GO decay responses, which normally lead to recycling of 40S and 60S subunits, and degradation of the mRNA and arrested nascent proteins. Overall, this study helps uncover specialized mechanisms in neurons that enable local protein translation at distal sites, supporting proper protein homeostasis, both crucial for neurodevelopment and optimal neuronal function.</jats:p>