CALR

CALR encodes calreticulin, an endoplasmic reticulum-resident protein in which recurrent somatic +1 frameshift mutations in exon 9 (most commonly a 52-bp deletion or 5-bp insertion) generate a novel positively-charged C-terminus while preserving ER retention (PMID:24325359). This mutant protein acquires a pathogenic gain of function: it preferentially binds the thrombopoietin receptor c-MPL complexed with JAK2, driving JAK2 phosphorylation and cytokine-independent proliferation, with the mutant-specific C-terminus disrupting the CALR P-domain so the N-domain engages c-MPL (PMID:26817954). This MPL-dependent activation of JAK-STAT signalling underlies mutant CALR-driven megakaryopoiesis and thrombocytosis in vivo, as demonstrated by mpl-knockdown epistasis in zebrafish and by knock-in mouse models that recapitulate an essential thrombocythemia-like phenotype (PMID:27716741, PMID:33761144). Receptor engagement requires both loss of the native KDEL motif and the positively-charged residues yet is not fully explained by them alone, since a murine del19 mutant binds MPL weakly and produces only mild disease (PMID:30926777). The neoepitope is therapeutically actionable: a monoclonal antibody against the mutant C-terminus selectively disrupts CALR-dimer binding to TpoR, blocking signalling, proliferation, and differentiation (PMID:35156745). Distinct from the classical chaperone calcium-buffering role, type 1 (del52) mutations impair the C-terminal calcium-binding activity more severely than type 2 (ins5), correlating with altered cytosolic calcium signalling (PMID:26449662).

1. 2013 High

   Established that recurrent exon 9 +1 frameshift mutations define a molecular class of myeloproliferative neoplasm, producing a mutant calreticulin with a novel C-terminus that nonetheless remains ER-retained.

   Evidence Exome sequencing with immunofluorescence and flow cytometry in a patient discovery cohort

   PMID:24325359

   Open questions at the time

   • Did not define the gain-of-function mechanism
   • Did not identify the receptor partner
2. 2015 Medium

   Addressed whether mutation subtypes differ biochemically, showing type 1 mutations impair C-terminal calcium binding more severely than type 2 and alter megakaryocyte calcium signalling.

   Evidence Calcium flux measurements in cultured megakaryocytes with isoelectric point prediction and clinical correlation

   PMID:26449662

   Open questions at the time

   • Single lab
   • Causal link between altered calcium and disease phenotype not established
   • Mechanism connecting calcium dysregulation to MPL signalling unresolved
3. 2016 High

   Identified the oncogenic gain-of-function: mutant CALR specifically binds the JAK2-bound thrombopoietin receptor c-MPL through its N-domain to drive cytokine-independent growth.

   Evidence Co-immunoprecipitation, cytokine-independent growth assays in UT-7/TPO cells, JAK2 inhibition, iPSC differentiation, and domain mutagenesis

   PMID:26817954

   Open questions at the time

   • Stoichiometry and structural basis of CALR-MPL engagement not resolved
   • Did not establish requirement in vivo
4. 2016 High

   Demonstrated in vivo genetic dependence on MPL, showing mutant CALR-induced thrombocytosis and JAK-STAT activation require mpl but not other cytokine receptors.

   Evidence Zebrafish transgenic expression with morpholino knockdown epistasis (mpl vs epor vs csf3r) and JAK inhibitor treatment

   PMID:27716741

   Open questions at the time

   • Zebrafish hematopoiesis may differ from mammalian
   • Did not address chronic disease evolution
5. 2019 Medium

   Clarified that loss of KDEL and acquisition of positively-charged C-terminal residues are necessary but not sufficient for full MPL binding, refining the structural requirements for oncogenic activity.

   Evidence CRISPR/Cas9 knock-in mouse (Calr del19) with in vitro binding and STAT5 phosphorylation assays

   PMID:30926777

   Open questions at the time

   • Species differences confound interpretation
   • Exact sequence features required for high-affinity MPL binding not defined
6. 2021 Medium

   Confirmed causality in human cells and an authentic in vivo disease model, with isogenic CRISPR correction and a conditional knock-in mouse recapitulating essential thrombocythemia.

   Evidence Patient-derived iPSCs with CRISPR repair and directed megakaryocyte differentiation; conditional knock-in mouse with transplantation, antibody treatment, and HSC transcriptomics

   PMID:33761144 PMID:34678208

   Open questions at the time

   • Single-lab models
   • Hypoxia and other dysregulated pathways not mechanistically linked to MPL signalling
   • Stem cell proliferative advantage mechanism incompletely defined
7. 2022 High

   Validated the mutant neoepitope as a therapeutic target by showing an antibody that disrupts CALR-dimer binding to TpoR blocks signalling and tumor growth.

   Evidence Monoclonal antibody (4D7) with cell-based proliferation, JAK-STAT readouts, patient samples, and xenograft models

   PMID:35156745

   Open questions at the time

   • Did not resolve whether dimerization is obligatory for receptor activation in all mutant types
   • Clinical efficacy not addressed
8. 2024 Medium

   Distinguished the calcium-signalling consequences of CALR mutation from JAK2-V617F, showing elevated cytosolic calcium during store depletion without hyperactivation of PLCγ-1 or IP3R.

   Evidence Fura-2 calcium measurements, SOCE assays, and phospho-western blotting in 32D myeloid progenitor cells

   PMID:38509561

   Open questions at the time

   • Mechanism producing altered calcium dynamics not identified
   • Functional contribution of calcium changes to myeloproliferation unclear
   • Single lab

• The structural basis of mutant CALR-MPL complex assembly and the contribution of calcium dysregulation to disease phenotype remain unresolved.
• No atomic-resolution structure of the mutant CALR-MPL-JAK2 complex
• Causal link between C-terminal calcium-binding loss and clinical phenotype not established
• Mechanism connecting altered cytosolic calcium to JAK-STAT output unknown