Whether protein translation occurs in the nucleus is contentious. To address this question, we developed the ribopuromycylation method (RPM), which visualizes translation in cells via standard immunofluorescence microscopy. The RPM is based on ribosome-catalyzed puromycylation of nascent chains immobilized on ribosomes by antibiotic chain elongation inhibitors followed by detection of puromycylated ribosome-bound nascent chains with a puromycin (PMY)-specific monoclonal antibody in fixed and permeabilized cells. The RPM correlates localized translation with myriad processes in cells and can be applied to any cell whose translation is sensitive to PMY. In this paper, we use the RPM to provide evidence for translation in the nucleoplasm and nucleolus, which is regulated by infectious and chemical stress.

https://rupress.org/jcb/article-pdf/197/1/45/1355857/jcb_201112145.pdf

Nuclear translation visualized by ribosome-bound nascent chain puromycylation

The structure and formation of ribosomes in animal cells

MHC class I molecules display oligopeptides on the cell surface to enable T cell immunosurveillance of intracellular pathogens and tumors. Speed is of the essence in detecting viruses, which can complete a full replication cycle in just hours, whereas tumor detection is typically a finding-the-needle-in-the-haystack exercise. We review current evidence supporting a nonrandom, compartmentalized selection of peptidogenic substrates that focuses on rapidly degraded translation products as a main source of peptide precursors to optimize immunosurveillance of pathogens and tumors.

https://jlb.onlinelibrary.wiley.com/doi/pdf/10.1189/jlb.1113599

Translating DRiPs: MHC class I immunosurveillance of pathogens and tumors

A General Survey of Mechanisms Regulating Protein Metabolism in Mammals

We have measured the cytokinetic changes which characterize the growth of a hyperdiploid Ehrlich ascites tumor and have investigated some possible underlying biochemical mechanisms. In this tumor, the cell cycle increases from 9.5 hr on the 2nd day of growth to 39 hr on the 14th day and the proportion of cells in S (thymidine index) declines from 80 to 40% during the same interval of growth. Protein synthesis (measured by uptake of leucine-14C during a 20-min pulse) declines at a rate similar to the decline of the thymidine index, but there are no changes in the specific activity of six enzymes: aryl sulfatase, β-glucuronidase, thymidine kinase, glucose-6-phosphate dehydrogenase, alkaline phosphatase, and glutathione reductase. Transplantation of 107 cells from plateau-phase tumors (14 to 16 days) to new hosts causes the rate of protein synthesis to increase immediately. The thymidine index, on the other hand, remains constant for 5 to 6 hr after transplantation and then doubles within the next 4 hr. The rise is prevented (or delayed) when the animals breathe 6% oxygen. Hypoxia also decreases the thymidine index of 2-day tumors, but it does not alter the length of the cell cycle. The results suggest that the progressive accumulation of cells in G1 during tumor growth may reflect the relative inefficiency of anaerobic metabolism in producing the energy ( e.g. , ATP) necessary for movement into S phase.

/pdf/biochemical-studies-of-cytokinetic-changes-during-tumor-j4ib5p5ble.pdf

Irwin M. Arias

Papers

Protein synthesis in isolated nuclei and nucleoli of HeLa cells.