iTRAQ proteomics dataset on ceramide-dependent exosomal cargoes from SW480 and SW620 cells

Cancer metastasis is largely influenced by cell–cell communication, to which exosomes play a vital role. Exosomes are small extracellular vesicles (sEVs) that originate as intraluminal vesicles (ILVs) within multivesicular bodies (MVBs) during endosome maturation. ILV formation depends on several pathways, including that of ceramide synthesis by neutral sphingomyelinase 2 [nSMase2]. Colorectal cancer (CRC)-derived sEVs are reported to carry a diverse range of metastatic cargo proteins; however, segregation of them in the ceramide-dependent sEV pool (sEVCer) remains unexplored. The current study aimed to identify the metastatic proteins that are secreted through sEVCer, from CRC cells of variable metastatic potentials. Primary (SW480) and metastatic (SW620) CRC cells were treated with nSMase2 blocker and sEVs were isolated, followed by extraction of the sEV proteins for a quantitative proteomic profiling using isobaric tags for relative and absolute quantitation (iTRAQ). In total, 1781 p..., The dataset was generated using 2D LC followed by tandem MS.
The tandem MS analysis was performed using a 5600 Triple TOF system (SCIEX) under Information Dependent Mode. The mass range of 400–1800 m/z and accumulation times of 250 ms per spectrum were chosen for precursor ion selections. MS/MS spectra were recorded in high sensitivity mode (resolution≥ 15 000) with rolling collision energy. In each cycle, a maximum of 20 precursors were selected for fragmentation.
Peptide and Protein Identification
Protein identification and relative iTRAQ quantification were performed with ProteinPilot™ Software 5.0 (AB SCIEX, revision number 4769) using the Paragon™ Algorithm (5.0.0.0, 4767) for the peptide identification, which was further processed by Pro GroupTM algorithm where isoform-specific quantification was adopted to trace the differences between expressions of various isoforms. The Pro Group Algorithm calculates protein ratios using only ratios from the spectra that are distinct to each pr..., , # iTRAQ proteomics dataset on ceramide-dependent exosomal cargoes from SW480 and SW620 cells

[https://doi.org/10.5061/dryad.r4xgxd2q8](https://doi.org/10.5061/dryad.r4xgxd2q8)

## Description of the data and file structure

Blocking the ceramide-dependent sEV biogenesis in SW480 and SW620 cells showed significant reduction in the quantity of released sEVs. As a consequence, the metastatic cargoes that are preferentially secreted through the ceramide-dependent route, are expected to be depleted in their sEV protein levels. These depleted cargoes are named as “Cargo-Cer” by us and will be referred as the same throughout the article. We hypothesized that the Cargo-Cer can vary with respect to the CRC stages, while some of them might remain overlapped. To specifically identify the Cargo-Cer for SW480 and SW620, we then performed iTRAQ-based quantitative proteomics with the proteins extracted from the sEVs of the respective cells. The mentioned cells were treated with GW4869, to block nSMas...