Project description:Ribosome profiling is a widespread tool for studying translational dynamics in human cells. Its central assumption is that ribosome footprint density on a transcript quantitatively reflects protein synthesis. Here, we test this assumption using pulsed-SILAC (pSILAC) high-accuracy targeted proteomics. We focus on multiple myeloma cells exposed to bortezomib, a first-line chemotherapy and proteasome inhibitor. In the absence of drug effects, we found that direct measurement of protein synthesis by pSILAC correlated well with indirect measurement of synthesis from ribosome footprint density. This correlation, however, broke down under bortezomib-induced stress. By developing a statistical model integrating longitudinal proteomic and mRNA-seq measurements, we found that proteomics could directly detect global alterations in translational rate caused by bortezomib; these changes are not detectable by ribosomal profiling alone. Further, by incorporating pSILAC data into a gene expression model, we predict cell-stress specific proteome remodeling events. These results demonstrate that pSILAC provides an important complement to ribosome profiling in measuring proteome dynamics. Timecourse experiment with six points over 48hr after bortezomib exposure in MM.1S myeloma cells. mRNA-seq and ribosome profiling data at each time point.
Project description:Oxaliplatin as a first-line drug frequently causes the chemo-resistance on colorectal cancer (CRC). N6-methyladenosine (m6A) methylation has been largely acknowledged in multiple biological functions. However, the molecular mechanisms underlying the m6A methylation in modulating anticancer drug resistance in CRC are still obscure. In present study, RNA-seq was conducted to investigate the transcriptome of HCT116, HCT116 cells with oxaliplatin resistance (HCT116R), HCT8 and HCT8 cells with oxaliplatin resistance (HCT8R).
Project description:Oxaliplatin as a first-line drug frequently causes the chemo-resistance on colorectal cancer (CRC). N6-methyladenosine (m6A) methylation has been largely acknowledged in multiple biological functions. However, the molecular mechanisms underlying the m6A methylation in modulating anticancer drug resistance in CRC are still obscure. In present study, RNA-seq was conducted to investigate the transcriptome of CRC tissues from three patients at different disease stages (CapeOx combined chemotherapy sensitivity and resistance).
Project description:Ribosome profiling is a widespread tool for studying translational dynamics in human cells. Its central assumption is that ribosome footprint density on a transcript quantitatively reflects protein synthesis. Here, we test this assumption using pulsed-SILAC (pSILAC) high-accuracy targeted proteomics. We focus on multiple myeloma cells exposed to bortezomib, a first-line chemotherapy and proteasome inhibitor. In the absence of drug effects, we found that direct measurement of protein synthesis by pSILAC correlated well with indirect measurement of synthesis from ribosome footprint density. This correlation, however, broke down under bortezomib-induced stress. By developing a statistical model integrating longitudinal proteomic and mRNA-seq measurements, we found that proteomics could directly detect global alterations in translational rate caused by bortezomib; these changes are not detectable by ribosomal profiling alone. Further, by incorporating pSILAC data into a gene expression model, we predict cell-stress specific proteome remodeling events. These results demonstrate that pSILAC provides an important complement to ribosome profiling in measuring proteome dynamics.