Ellinor Johnsson

Variants in ELL2 influencing immunoglobulin levels associate with multiple myeloma

Multiple myeloma (MM) is characterized by an uninhibited, clonal growth of plasma cells. While first-degree relatives of patients with MM show an increased risk of MM, the genetic basis of inherited MM susceptibility is incompletely understood. Here we report a genome-wide association study in the Nordic region identifying a novel MM risk locus at ELL2 (rs56219066T; odds ratio (OR)=1.25; P=9.6 × 10−10). This gene encodes a stoichiometrically limiting component of the super-elongation complex that drives secretory-specific immunoglobulin mRNA production and transcriptional regulation in plasma cells. We find that the MM risk allele harbours a Thr298Ala missense variant in an ELL2 domain required for transcription elongation. Consistent with a hypomorphic effect, we find that the MM risk allele also associates with reduced levels of immunoglobulin A (IgA) and G (IgG) in healthy subjects (P=8.6 × 10−9 and P=6.4 × 10−3, respectively) and, potentially, with an increased risk of bacterial meningitis (OR=1.30; P=0.0024).

The Complete Mitochondrial DNA Sequence of the Harbor Seal, Phoca vitulina

SummaryThe nucleotide sequence of the mitochondrial DNA (mtDNA) of the harbor seal, Phoca vitulina, was determined. The total length of the molecule was 16,826 bp. The organization of the coding regions of the molecule conforms with that of other mammals, but the control region is unusually long. A considerable portion of the control region is made up of short repeats with the motif GTACAC particularly frequent. The two rRNA genes and the 13 peptide-coding genes of the harbor seal, fin whale, cow, human, mouse, and rat were compared and the relationships between the different species assessed. At ordinal level the 12S rRNA gene and 7 out of the 13 peptide-coding genes yielded a congruent topological tree of the mtDNA relationship between the seal, cow, whale, human, and the rodents. In this tree the whale and the cow join first, and this clade is most closely related to the seal.

The nucleotide sequence of the mitochondrial DNA molecule of the grey seal, Halichoerus grypus, and a comparison with mitochondrial sequences of other true seals

The sequence of the mtDNA of the grey seal, Halichoerus grypus, was determined. The length of the molecule was 16,797 base pairs. The organization of the molecule conformed with that of other eutherian mammals but the control region was unusually long due to the presence of two types of repeated motifs. The grey seal and the previously reported harbor seal, Phoca vitulina, belong to different but closely related genera of family Phocidae, true (or earless) seals. In order to determine the degree of differences that may occur between mtDNAs of closely related mammalian genera, the 2 rRNA genes, the 13 peptide coding genes, and the 22 tRNA genes of the 2 species were compared. Total nucleotide difference in the peptide coding genes was 2.0–6.1%. The range of conservative difference was 0.0–1.5%. In the inferred peptide sequences the amino acid difference was 0.0–4.5%, and the difference with respect to chemical properties of amino acids was 0.0–3.0%. A gene that showed a limited degree of difference in one mode of comparison did not necessarily show a corresponding limited difference in another mode. The ratio for differences in codon positions 1, 2, and 3 was ≈2.7:1:16. The corresponding ratio for conservative differences was ≈ 1.8:1. l:1. The evolutionary separation of the two species was calculated to have taken place 2–2.5 million years ago. This dating gives the figure ≈8 × 10−9 as the mean rate of substitution per site and year in the entire mtDNA molecule. Comparison with the cytochrome b gene of the Hawaiian monk seal and the Weddell seal suggested that the lineage of these two species and that of the grey and harbor seals separated ≈8 million years ago.

Genome-wide association study identifies multiple susceptibility loci for multiple myeloma

Multiple myeloma (MM) is a plasma cell malignancy with a significant heritable basis. Genome-wide association studies have transformed our understanding of MM predisposition, but individual studies have had limited power to discover risk loci. Here we perform a meta-analysis of these GWAS, add a new GWAS and perform replication analyses resulting in 9,866 cases and 239,188 controls. We confirm all nine known risk loci and discover eight new loci at 6p22.3 (rs34229995, P=1.31 × 10−8), 6q21 (rs9372120, P=9.09 × 10−15), 7q36.1 (rs7781265, P=9.71 × 10−9), 8q24.21 (rs1948915, P=4.20 × 10−11), 9p21.3 (rs2811710, P=1.72 × 10−13), 10p12.1 (rs2790457, P=1.77 × 10−8), 16q23.1 (rs7193541, P=5.00 × 10−12) and 20q13.13 (rs6066835, P=1.36 × 10−13), which localize in or near to JARID2, ATG5, SMARCD3, CCAT1, CDKN2A, WAC, RFWD3 and PREX1. These findings provide additional support for a polygenic model of MM and insight into the biological basis of tumour development.

Robust isolation of malignant plasma cells in multiple myeloma

Molecular characterization of malignant plasma cells is increasingly important for diagnostic and therapeutic stratification in multiple myeloma. However, the malignant plasma cells represent a relatively small subset of bone marrow cells, and need to be enriched prior to analysis. Currently, the cell surface marker CD138 (SDC1) is used for this enrichment, but has an important limitation in that its expression decreases rapidly after sampling. Seeking alternatives to CD138, we performed a computational screen for myeloma plasma cell markers and systematically evaluated 7 candidates. Our results conclusively show that the markers CD319 (SLAMF7/CS1) and CD269 (TNFRSF17/BCMA) are considerably more robust than CD138 and enable isolation of myeloma plasma cells under more diverse conditions, including the samples that have been delayed or frozen. Our results form the basis of improved procedures for characterizing cases of multiple myeloma in clinical practice.

Identification of sequence variants influencing immunoglobulin levels

Immunoglobulins are the effector molecules of the adaptive humoral immune system. In a genome-wide association study of 19,219 individuals, we found 38 new variants and replicated 5 known variants associating with IgA, IgG or IgM levels or with composite immunoglobulin traits, accounted for by 32 loci. Variants at these loci also affect the risk of autoimmune diseases and blood malignancies and influence blood cell development. Notable associations include a rare variant at RUNX3 decreasing IgA levels by shifting isoform proportions (rs188468174[C>T]: P = 8.3 × 10−55, β = −0.90 s.d.), a rare in-frame deletion in FCGR2B abolishing IgG binding to the encoded receptor (p.Asn106del: P = 4.2 × 10−8, β = 1.03 s.d.), four IGH locus variants influencing class switching, and ten new associations with the HLA region. Our results provide new insight into the regulation of humoral immunity.

Artificially controlled aggregation of proteins and targeting in hematopoietic cells

The targeting mechanisms for granule proteins in hematopoietic cells are largely unknown. Aggregation is believed to be important for protein sorting‐for‐entry and sorting‐by‐retention in endocrine and neuroendocrine cells. We asked whether artificially induced multimerization/aggregation of chimeric proteins could affect their sorting in hematopoietic cells. A system was used that permits ligand‐controlled intracellular oligomerization of hybrid proteins containing the FK506‐binding protein (FKBP). The hybrid proteins ELA‐(FKBP)3 with neutrophil elastase (ELA) and (FKBP∗)4‐FCS‐hGH with a furin cleavage site (FCS) and human growth hormone (hGH) were expressed in the myeloblastic 32D and the rat basophilic leukemia (RBL‐1) hematopoietic cell lines. ELA alone is normally targeted to secretory lysosomes. However, the hybrid proteins and ligand‐induced aggregates of them were constitutively secreted and not targeted. The hGH that was released at the FCS in (FKBP∗)4‐FCS‐hGH was also constitutively secreted. We conclude that protein multimerization/aggregation per se is not enough to facilitate sorting‐for‐entry to secretory lysosomes in hematopoietic cells and that improperly folded proteins may be eliminated from sorting by constitutive secretion.

Direct evidence for a polygenic etiology in familial multiple myeloma

Although common risk alleles for multiple myeloma (MM) were recently identified, their contribution to familial MM is unknown. Analyzing 38 familial cases identified primarily by linking Swedish nationwide registries, we demonstrate an enrichment of common MM risk alleles in familial compared with 1530 sporadic cases (P = 4.8 × 10-2 and 6.0 × 10-2, respectively, for 2 different polygenic risk scores) and 10 171 population-based controls (P = 1.5 × 10-4 and 1.3 × 10-4, respectively). Using mixture modeling, we estimate that about one-third of familial cases result from such enrichments. Our results provide the first direct evidence for a polygenic etiology in a familial hematologic malignancy.

The multiple myeloma risk allele at 5q15 lowers ELL2 expression and increases ribosomal gene expression

Recently, we identified ELL2 as a susceptibility gene for multiple myeloma (MM). To understand its mechanism of action, we performed expression quantitative trait locus analysis in CD138+ plasma cells from 1630 MM patients from four populations. We show that the MM risk allele lowers ELL2 expression in these cells (Pcombined = 2.5 × 10−27; βcombined = −0.24 SD), but not in peripheral blood or other tissues. Consistent with this, several variants representing the MM risk allele map to regulatory genomic regions, and three yield reduced transcriptional activity in plasmocytoma cell lines. One of these (rs3777189-C) co-locates with the best-supported lead variants for ELL2 expression and MM risk, and reduces binding of MAFF/G/K family transcription factors. Moreover, further analysis reveals that the MM risk allele associates with upregulation of gene sets related to ribosome biogenesis, and knockout/knockdown and rescue experiments in plasmocytoma cell lines support a cause–effect relationship. Our results provide mechanistic insight into MM predisposition.ELL2 was recently discovered as a susceptibility gene for multiple myeloma (MM). Here, they show that the MM risk allele lowers ELL2 expression in plasma cells, that it also upregulates gene sets related to ribosome biogenesis, and that one of the linked variants reduces binding of MAFF/G/K family transcription factors.