Defining the cancer-specific proteome in Calreticulin-mutant myeloproliferative neoplasms

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Philadelphia-negative myeloproliferative neoplasms comprise a group of malignant hematological diseases. The genetic landscape of myeloproliferative neoplasms is very well described, with calreticulin being the second most common mutated gene after JAK2. Calreticulin mutations are found in up to 40% of patients with myeloproliferative neoplasms. Moreover, calreticulin and JAK2V617F mutations are mutually exclusive. All calreticulin mutations cluster in the last coding exon and result in deletion of the C-terminal “KDEL“ sequence, which normally retains wildtype calreticulin to the endoplasmic reticulum where it acts as a chaperone protein. The two most prominent mutations (Type I and II) both lead to frameshifts that generate a common mutant-specific 36-amino acid C-terminal peptide. In the new C-terminus, negatively charged amino acids are replaced by positively charged ones, mainly arginine and lysine. Recently, one mechanism by which mutant calreticulin is oncogenic has been delineated. Dr. Mullally’s lab showed that mutant calreticulin develops a pathogenic interaction with the thrombopoietin receptor, which activates the downstream signaling pathway in a thrombopoietin-independent manner. Moreover, it has been demonstrated that both the positive electrostatic charge of the mutant calreticulin C-terminus and the lectin binding residues of mutant calreticulin are required for its oncogenic activity. In work proposed here, I will characterise the role of novel interaction partners of mutant calreticulin in the development of myeloproliferative neoplasms. Moreover, I will determine how mutant calreticulin aberrantly binds to its partners and assess the necessity of this binding in mutant calreticulin cell proliferation. Dr. Mullaly’s group has also shown that mutant calreticulin loses its chaperone activity. In a second approach, I will therefore determine whether mutant calreticulin induces proteome variations such as aberrantly folded proteins at the cell surface. These non-canonical proteins would, by definition, only be present on calreticulin mutated, malignant cells and therefore constitute ideal targets of future immunological therapy. I anticipate identifying non-canonical protein isoforms due to the aberrant chaperone activity of mutant calreticulin. Candidate isoforms will be confirmed and subsequently validated in primary cells.