👤 Frederic Sorgeloos

Proteins of pathogens such as cardioviruses, Kaposi sarcoma-associated herpes virus, varicella zoster virus and bacteria of the genus Yersinia were previously shown to use a common "DDVF" (D/E-D/E-V-F) short linear motif (SLiM) to hijack cellular kinases of the RSK (p90 ribosomal S6 kinases) family. Notably, the leader (L) protein of Theiler's murine encephalomyelitis virus (TMEV), a cardiovirus, and protein YopM of Yersinia species were shown to act as adapters to retarget RSKs toward unconventional substrates, nucleoporins and pyrin, respectively. Remarkable conservation of the SLiM docking site targeted by pathogens' proteins in RSK sequences suggested a physiological role for this site. Using SLiM prediction tools and AlphaFold docking, we screened the human proteome for proteins that would interact with RSKs through a DDVF-like SLiM. Co-immunoprecipitation experiments show that two candidates previously known as RSK partners, FGFR1 and SPRED2, as well as two candidates identified as novel RSK partners, GAB3 and CNKSR2 do interact with RSKs through a similar interface as the one used by pathogens, as was recently documented for SPRED2. FGFR1 employs a DSVF motif to bind RSKs and phosphorylation of the serine in this motif slightly increased RSK binding. FGFR1, SPRED2, GAB3 and CNKSR2 act upstream of RSK in the RAS-ERK MAP kinase pathway. Analysis of ERK activation in cells expressing a mutated form of RSK lacking the DDVF-docking site suggests that RSK might interact with the DDVF-like SLiM of several partners to provide a negative feed-back to the ERK MAPK pathway. Moreover, after TMEV infection, ERK phosphorylation was altered by the L protein in a DDVF-dependent manner. Taken together, our data suggest that, in addition to retargeting RSKs toward unconventional substrates, pathogens' proteins carrying a DDVF-like motif can compete with endogenous DDVF-containing proteins for RSK binding, thereby altering the regulation of the RAS-ERK MAP kinase pathway. Show less

This study was designed to identify significant differences in gene expression profiles of human papillomavirus (HPV)-positive and HPV-negative oropharyngeal squamous cell carcinomas (OPSCC) and to better understand the functional and biological effects of HPV infection in the premalignant pathway. Twenty-four consecutive patients with locally advanced primary OPSCC were included in a prospective clinical trial. Fresh tissue samples (tumor vs. matched normal epithelium) were subjected to whole transcriptome analysis and the results validated on the same cohort with RT-quantitative real-time PCR. In a separate retrospective cohort of 27 OPSCC patients, laser capture microdissection of formalin-fixed, paraffin-embedded tissue allowed RNA extraction from adjacent regions of normal epithelium, carcinoma in situ (premalignant) and invasive SCC tissue. The majority of patients showed evidence of high-risk HPV16 positivity (80.4%). Predictable fold changes of RNA expression in HPV-associated disease included multiple transcripts within the p53 oncogenic pathway (e.g. CDKN2A/CCND1). Other candidate transcripts found to have altered levels of expression in this study have not previously been established (SFRP1, CRCT1, DLG2, SYCP2, and CRNN). Of these, SYCP2 showed the most consistent fold change from baseline in premalignant tissue; aberrant expression of this protein may contribute to genetic instability during HPV-associated cancer development. If further corroborated, this data may contribute to the development of a non-invasive screening tool. This study is registered with the UK Clinical Research Network (ref.: 11945). Show less