Gamma Herpesviruses Modulate Exosome Proteins

Exosomes are small microvescicle compartments that originate from internal endosomal-derived membranes of cells. After exosomes are released from a cell, they can travel through bodily fluids and be taken up by neighboring cells or travel to cells in other areas of the body. Exosomes play a role in diseases pathologies, such as the human gamma herpesviruses, Kaposi’s sarcoma-associated herpesvirus (KSHV) and the Epstein–Barr virus (EBV), and are associated with multiple types of lymphomas. EBV-infected cells have been shown to transfer virally modiﬁed exosomes and modulate the environment and function of cells. Researchers have also identified oncoproteins latent membrane proteins 1 and 2 (LMP1 and LMP2) and virally encoded microRNAs (miRNAs) present in EBV-modified exosomes.¹Mounting evidence suggests that EBV and KSHV can also contribute to cancer progression through infected exosomes.²

A recent publication describes work by Meckes et al. (2013) that is aimed at identifying how EBV and KSHV infections alter the cellular component of exosomes.² The researchers utilized a proteomics strategy to analyze puriﬁed exosomes from EBV-infected, KSHV-infected and dually infected cells derived from B-cell lines. For their experiments, proteins were extracted and digested, and puriﬁed exosome proteins were separated using strong anion-exchange and reverse-phase chromatography, utilizing an Easy Nano Liquid Chromatography II system (Thermo Scientiﬁc). The liquid chromatography system was coupled to an LTQ Velos linear ion trap mass spectrometer (Thermo Scientific).

Analysis by mass spectrometry revealed a total of 871 non-viral proteins throughout all B-cell exosome lines. Five hundred sixty-nine proteins were present in the Exocarta database of published exosome proteomic results, including 21 of the 25 most frequently identiﬁed proteins; 302 proteins represented newly identiﬁed exosome components not present in Exocarta exosome database.

When researchers examined the protein content of EBV- or KSHV-infected samples, they found that 449 proteins were common in uninfected B-cell exosomes and 230 proteins were only present in virus-infected exosomes, with 93 proteins specific to EBV-infected exosomes and 22 proteins specific to KSHV-infected exosomes.

Protein quantification was performed using the Kyoto Encyclopedia of Genes and Genomes pathway analysis, along with a two-tailed ANOVA test functional categorization. As a result, 164 proteins had signiﬁcantly (p ≤ 0.05) altered levels due to virus infection, as compared with the uninfected control cell line speciﬁc to the KSHV exosomes. Sixteen distinct proteins were present in the dually infected cell lines. Interestingly, 2D DIGE and spectral counting revealed that dually infected exosomes had more in common with KSHV-infected exosomes than with EBV-infected exosomes, suggesting that EBV has less of an influence on exosomes. LMP1 was detected in EBV+ cell lines using immunoblotting but was not detected in the mass spectrometry analysis.

Changes between herpesvirus-infected exosomes and uninfected controls indicated that the altered exosomes do manipulate the cellular environment by modulating cell death, protein synthesis and various signaling pathways. Meckes et al. found these results encouraging and they predict additional intriguing findings as the exosome proteome is further explored.

References

1. Meckes, D.G., Jr., et al. (2010) “Human tumor virus utilizes exosomes for intercellular communication,” Proceedings of the National Academy of Sciences of the United States of America, 107(47) (pp. 20370–5).

2. Meckes, D.G., Jr., et al. (2013, July 30) “Modulation of B-cell exosome proteins by gamma herpesvirus infection,” Proceedings of the National Academy of Sciences of the United States of America, 110(31) (pp. E2925–33).

Post Author: Emily Humphreys. As a biology undergraduate at the University of Utah, Emily balanced a heavy class schedule while working long hours in a lab studying eye development. Following graduation, she became involved in infectious disease and aging research involving SNPS.

While she enjoyed the thrill of research, Emily has since traded bench work for science journalism.
And has been a regular contributor to Accelerating Science since 2012.