Kärt Tomberg | The Wellcome Trust Sanger Institute (original) (raw)

Papers by Kärt Tomberg

ABSTRACTThe COPII component SEC24 mediates the recruitment of transmembrane cargoes or cargo adap... more ABSTRACTThe COPII component SEC24 mediates the recruitment of transmembrane cargoes or cargo adaptors into newly forming COPII vesicles on the ER membrane. Mammalian genomes encode four Sec24 paralogs (Sec24a-d), with two subfamilies based on sequence homology (SEC24A/B and C/D), though little is known about their comparative functions and cargo-specificities. Complete deficiency for Sec24d results in very early embryonic lethality in mice (before the 8 cell stage), with later embryonic lethality (E 7.5) observed in Sec24c null mice. To test the potential overlap in function between SEC24C/D, we employed dual recombinase mediated cassette exchange to generate a Sec24cc-d allele, in which the C-terminal 90% of SEC24C has been replaced by SEC24D coding sequence. In contrast to the embryonic lethality at E7.5 of SEC24C-deficiency, Sec24cc-d/c-d pups survive to term, though dying shortly after birth. Sec24cc-d/c-d pups are smaller in size, but exhibit no obvious developmental abnormalit...

Proceedings of the National Academy of Sciences of the United States of America, Sep 5, 2017

Factor V Leiden (F5(L) ) is a common genetic risk factor for venous thromboembolism in humans. We... more Factor V Leiden (F5(L) ) is a common genetic risk factor for venous thromboembolism in humans. We conducted a sensitized N-ethyl-N-nitrosourea (ENU) mutagenesis screen for dominant thrombosuppressor genes based on perinatal lethal thrombosis in mice homozygous for F5(L) (F5(L/L) ) and haploinsufficient for tissue factor pathway inhibitor (Tfpi(+/-) ). F8 deficiency enhanced the survival of F5(L/L)Tfpi(+/-) mice, demonstrating that F5(L/L)Tfpi(+/-) lethality is genetically suppressible. ENU-mutagenized F5(L/L) males and F5(L/+)Tfpi(+/-) females were crossed to generate 6,729 progeny, with 98 F5(L/L)Tfpi(+/-) offspring surviving until weaning. Sixteen lines, referred to as "modifier of Factor 5 Leiden (MF5L1-16)," exhibited transmission of a putative thrombosuppressor to subsequent generations. Linkage analysis in MF5L6 identified a chromosome 3 locus containing the tissue factor gene (F3). Although no ENU-induced F3 mutation was identified, haploinsufficiency for F3 (F3(+/-...

Factor V Leiden (F5L) is a common genetic risk factor for venous thromboembolism in humans. We co... more Factor V Leiden (F5L) is a common genetic risk factor for venous thromboembolism in humans. We conducted a sensitized ENU mutagenesis screen for dominant thrombosuppressor genes based on perinatal lethal thrombosis in mice homozygous for F5L (F5L/L) and haploinsufficient for tissue factor pathway inhibitor (Tfpi+/-). F8 deficiency enhanced survival of F5L/L Tfpi+/- mice, demonstrating that F5L/L Tfpi+/- lethality is genetically suppressible. ENU-mutagenized F5L/L males and F5L/+ Tfpi+/- females were crossed to generate 6,729 progeny, with 98 F5L/L Tfpi+/- offspring surviving until weaning. Sixteen lines exhibited transmission of a putative thrombosuppressor to subsequent generations, with these lines referred to as MF5L (Modifier of Factor 5 Leiden) 1-16. Linkage analysis in MF5L6 identified a chromosome 3 locus containing the tissue factor gene (F3). Though no ENU-induced F3 mutation was identified, haploinsufficiency for F3 (F3+/-) suppressed F5L/L Tfpi+/- lethality. Whole exome s...

BMC Cardiovascular Disorders, 2011

Background: More than one third of adult population in Estonia has problems with elevated blood p... more Background: More than one third of adult population in Estonia has problems with elevated blood pressure (BP). The Hypertension in Estonia (HYPEST) study represents the country's first hypertension-targeted sample collection aiming to examine the epidemiological and genetic determinants for hypertension (HTN) and related cardiovascular diseases (CVD) in Estonian population. The HYPEST subjects (n = 1,966) were recruited across Estonia between 2004-2007 including clinically diagnosed HTN cases and population-based controls. The present report is focused on the clinical and epidemiological profile of HYPEST cases, and gender-specific effects on the pathophysiology of hypertension. Methods: Current analysis was performed on 1,007 clinically diagnosed HTN patients (617 women and 390 men) aged 18-85 years. The hypertensives were recruited to the study by BP specialists at the North Estonia Medical Center, Centre of Cardiology, Tallinn or at the Cardiology Clinic, Tartu University Hospital, Estonia. Longitudinal BP data was extracted retrospectively from clinical records. Current and retrospective data of patient's medical history, medication intake and lifestyle habits were derived from self-administrated questionnaire and each variable was examined separately for men and women. Eleven biochemical parameters were measured from fasting serum samples of 756 patients. Results: The distribution of recruited men and women was 39% and 61% respectively. Majority of Estonian HTN patients (85%) were overweight (BMI ≥ 25 kg/m 2 ) and a total of 79% of patients had additional complications with cardiovascular system. In men, the hypertension started almost 5 years earlier than in women (40.5 ± 14.5 vs 46.1 ± 12.7 years), which led to earlier age of first myocardial infarction (MI) and overall higher incidence rate of MI among male patients (men 21.2%, women 8.9%, P < 0.0001). Heart arrhythmia, thyroid diseases, renal tubulointestinal diseases and hyperlipidemia were more prevalent in hypertensive women compared to men (P < 0.0001). An earlier age of HTN onset was significantly associated with smoking (P = 0.00007), obesity (BMI ≥ 30 kg/m 2 ; P = 0.0003), increased stress (P = 0.0003) and alcohol consumption (P = 0.004).

Nature Biotechnology, 2018

Background: More than one third of adult population in Estonia has problems with elevated blood p... more Background: More than one third of adult population in Estonia has problems with elevated blood pressure (BP).
The Hypertension in Estonia (HYPEST) study represents the country’s first hypertension-targeted sample collection
aiming to examine the epidemiological and genetic determinants for hypertension (HTN) and related
cardiovascular diseases (CVD) in Estonian population. The HYPEST subjects (n = 1,966) were recruited across Estonia
between 2004-2007 including clinically diagnosed HTN cases and population-based controls. The present report is
focused on the clinical and epidemiological profile of HYPEST cases, and gender-specific effects on the
pathophysiology of hypertension.
Methods: Current analysis was performed on 1,007 clinically diagnosed HTN patients (617 women and 390 men) aged
18-85 years. The hypertensives were recruited to the study by BP specialists at the North Estonia Medical Center, Centre
of Cardiology, Tallinn or at the Cardiology Clinic, Tartu University Hospital, Estonia. Longitudinal BP data was extracted
retrospectively from clinical records. Current and retrospective data of patient’s medical history, medication intake and
lifestyle habits were derived from self-administrated questionnaire and each variable was examined separately for men
and women. Eleven biochemical parameters were measured from fasting serum samples of 756 patients.
Results: The distribution of recruited men and women was 39% and 61% respectively. Majority of Estonian HTN
patients (85%) were overweight (BMI ≥ 25 kg/m2) and a total of 79% of patients had additional complications with
cardiovascular system. In men, the hypertension started almost 5 years earlier than in women (40.5 ± 14.5 vs 46.1 ±
12.7 years), which led to earlier age of first myocardial infarction (MI) and overall higher incidence rate of MI
among male patients (men 21.2%, women 8.9%, P < 0.0001). Heart arrhythmia, thyroid diseases, renal tubulointestinal
diseases and hyperlipidemia were more prevalent in hypertensive women compared to men (P < 0.0001).
An earlier age of HTN onset was significantly associated with smoking (P = 0.00007), obesity (BMI ≥ 30 kg/m2; P =
0.0003), increased stress (P = 0.0003) and alcohol consumption (P = 0.004).
Conclusion: Understanding the clinical profile of HTN patients contributes to CVD management. Estonian
hypertension patients exhibited different disease and risk profiles of male and female patients. This wellcharacterized
sample set provides a good resource for studying hypertension and other cardiovascular phenotypes.

Introduction: Recurrent miscarriage (RM; 3 consecutive pregnancy losses) occurs in 1e3% of ferti... more Introduction: Recurrent miscarriage (RM; 3 consecutive pregnancy losses) occurs in 1e3% of fertile
couples. No biomarkers with high predictive value of threatening miscarriage have been identified. We
aimed to profile whole-genome differential gene expression in RM placental tissue, and to determine the
protein levels of identified loci in maternal sera in early pregnancy.
Methods: GeneChips (Affymetrix) were used for discovery and Taqman RT-qPCR assays for replication of
mRNA expression in placentas from RM cases (n ¼ 13) compared to uncomplicated pregnancies matched
for gestational age (n ¼ 23). Concentrations of soluble TRAIL (sTRAIL) and calprotectin in maternal serum
in normal first trimester (n ¼ 35) and failed pregnancies (early miscarriage, n ¼ 18, late miscarriage,
n ¼ 4; tubal pregnancy, n ¼ 11) were determined using ELISA.
Results: In RM placentas 30 differentially expressed (with nominal P-value < 0.05) transcripts were
identified. Significantly increased placental mRNA expression of TNF-related apoptosis-inducing ligand
(TRAIL; P ¼ 1.4 103; fold-change 1.68) and S100A8 (P ¼ 7.9 104; fold-change 2.56) encoding for
inflammatory marker calprotectin (S100A8/A9) was confirmed by RT-qPCR. When compared to normal
first trimester pregnancy (sTRAIL 16.1 1.6 pg/ml), significantly higher maternal serum concentration of
sTRAIL was detected at the RM event (33.6 4.3 pg/ml, P ¼ 0.00027), and in pregnant women, who
developed an unpredicted miscarriage 2e50 days after prospective serum sampling (28.5 4.4 pg/ml,
P ¼ 0.039). Women with tubal pregnancy also exhibited elevated sTRAIL (30.5 3.9 pg/ml, P ¼ 0.035).
Maternal serum levels of calprotectin were neither diagnostic nor prognostic to early pregnancy failures
(P > 0.05).
Conclusions: The study indicated of sTRAIL as a potential predictive biomarker in maternal serum for
early pregnancy complications.

Hypertension is a complex disease that affects a large proportion of adult population. Although a... more Hypertension is a complex disease that affects a large proportion of adult population. Although approximately
half of the inter-individual variance in blood pressure (BP) level is heritable, identification of genes
responsible for its regulation has remained challenging. Genome-wide association study (GWAS) is a
novel approach to search for genetic variants contributing to complex diseases. We conducted GWAS for
three BP traits [systolic and diastolic blood pressure (SBP and DBP); hypertension (HYP)] in the
Kooperative Gesundheitsforschung in der Region Augsburg (KORA) S3 cohort (n 5 1644) recruited from general
population in Southern Germany. GWAS with 395 912 single nucleotide polymorphisms (SNPs) identified
an association between BP traits and a common variant rs11646213 (T/A) upstream of the CDH13 gene at
16q23.3. The initial associations with HYP and DBP were confirmed in two other European populationbased
cohorts: KORA S4 (Germans) and HYPEST (Estonians). The associations between rs11646213 and three BP traits were replicated in combined analyses (dominant model: DBP, P 5 5.55 3 10–5, effect
–1.40 mmHg; SBP, P 5 0.007, effect –1.56 mmHg; HYP, P 5 5.30 3 1028, OR5 0.67). Carriers of the minor
allele A had a decreased risk of hypertension. A non-significant trend for association was also detected
with severe family based hypertension in the BRIGHT sample (British). The novel susceptibility locus,
CDH13, encodes for an adhesion glycoprotein T-cadherin, a regulator of vascular wall remodeling and angiogenesis.
Its function is compatible with the BP biology and may improve the understanding of the pathogenesis
of hypertension.

In humans, loss of function mutations in SEC23B result in Congenital Dyserythropoietic Anemia typ... more In humans, loss of function mutations in SEC23B result in Congenital Dyserythropoietic Anemia type
II (CDAII), a disease limited to defective erythroid development. Patients with two nonsense SEC23B
mutations have not been reported, suggesting that complete SEC23B deficiency might be lethal.
We previously reported that SEC23B-deficient mice die perinatally, exhibiting massive pancreatic
degeneration and that mice with hematopoietic SEC23B deficiency do not exhibit CDAII. We now
show that SEC23B deficiency restricted to the pancreas is sufficient to explain the lethality observed
in mice with global SEC23B-deficiency. Immunohistochemical stains demonstrate an acinar cell defect
but normal islet cells. Mammalian genomes contain two Sec23 paralogs, Sec23A and Sec23B. The
encoded proteins share ~85% amino acid sequence identity. We generate mice with pancreatic SEC23A
deficiency and demonstrate that these mice survive normally, exhibiting normal pancreatic weights and
histology. Taken together, these data demonstrate that SEC23B but not SEC23A is essential for murine
pancreatic development. We also demonstrate that two BAC transgenes spanning Sec23b rescue the
lethality of mice homozygous for a Sec23b gene trap allele, excluding a passenger gene mutation as
the cause of the pancreatic lethality, and indicating that the regulatory elements critical for Sec23b
pancreatic function reside within the BAC transgenes.

Proteases play important roles in many biologic processes and are key mediators of cancer, inflam... more Proteases play important roles in many biologic processes and are
key mediators of cancer, inflammation, and thrombosis. However,
comprehensive and quantitative techniques to define the substrate
specificity profile of proteases are lacking. The metalloprotease
ADAMTS13 regulates blood coagulation by cleaving vonWillebrand
factor (VWF), reducing its procoagulant activity. A mutagenized
substrate phage display library based on a 73-amino acid fragment
of VWF was constructed, and the ADAMTS13-dependent change in
library complexity was evaluated over reaction time points, using
high-throughput sequencing. Reaction rate constants (kcat/KM) were
calculated for nearly every possible single amino acid substitution
within this fragment. This massively parallel enzyme kinetics analysis
detailed the specificity of ADAMTS13 and demonstrated the
critical importance of the P1-P1′ substrate residues while defining
exosite binding domains. These data provided empirical evidence
for the propensity for epistasis within VWF and showed strong
correlation to conservation across orthologs, highlighting evolutionary
selective pressures for VWF.

PloS one, 2016

During the analysis of a whole genome ENU mutagenesis screen for thrombosis modifiers, a spontane... more During the analysis of a whole genome ENU mutagenesis screen for thrombosis modifiers, a spontaneous 8 base pair (bp) deletion causing a frameshift in exon 27 of the Nbeal2 gene was identified. Though initially considered as a plausible thrombosis modifier, this Nbeal2 mutation failed to suppress the synthetic lethal thrombosis on which the original ENU screen was based. Mutations in NBEAL2 cause Gray Platelet Syndrome (GPS), an autosomal recessive bleeding disorder characterized by macrothrombocytopenia and gray-appearing platelets due to lack of platelet alpha granules. Mice homozygous for the Nbeal2 8 bp deletion (Nbeal2gps/gps) exhibit a phenotype similar to human GPS, with significantly reduced platelet counts compared to littermate controls (p = 1.63 x 10-7). Nbeal2gps/gps mice also have markedly reduced numbers of platelet alpha granules and an increased level of emperipolesis, consistent with previously characterized mice carrying targeted Nbeal2 null alleles. These findings...

Proceedings of the National Academy of Sciences of the United States of America, Sep 5, 2017

BMC Cardiovascular Disorders, 2011

Nature Biotechnology, 2018

Background: More than one third of adult population in Estonia has problems with elevated blood p... more Background: More than one third of adult population in Estonia has problems with elevated blood pressure (BP).
The Hypertension in Estonia (HYPEST) study represents the country’s first hypertension-targeted sample collection
aiming to examine the epidemiological and genetic determinants for hypertension (HTN) and related
cardiovascular diseases (CVD) in Estonian population. The HYPEST subjects (n = 1,966) were recruited across Estonia
between 2004-2007 including clinically diagnosed HTN cases and population-based controls. The present report is
focused on the clinical and epidemiological profile of HYPEST cases, and gender-specific effects on the
pathophysiology of hypertension.
Methods: Current analysis was performed on 1,007 clinically diagnosed HTN patients (617 women and 390 men) aged
18-85 years. The hypertensives were recruited to the study by BP specialists at the North Estonia Medical Center, Centre
of Cardiology, Tallinn or at the Cardiology Clinic, Tartu University Hospital, Estonia. Longitudinal BP data was extracted
retrospectively from clinical records. Current and retrospective data of patient’s medical history, medication intake and
lifestyle habits were derived from self-administrated questionnaire and each variable was examined separately for men
and women. Eleven biochemical parameters were measured from fasting serum samples of 756 patients.
Results: The distribution of recruited men and women was 39% and 61% respectively. Majority of Estonian HTN
patients (85%) were overweight (BMI ≥ 25 kg/m2) and a total of 79% of patients had additional complications with
cardiovascular system. In men, the hypertension started almost 5 years earlier than in women (40.5 ± 14.5 vs 46.1 ±
12.7 years), which led to earlier age of first myocardial infarction (MI) and overall higher incidence rate of MI
among male patients (men 21.2%, women 8.9%, P < 0.0001). Heart arrhythmia, thyroid diseases, renal tubulointestinal
diseases and hyperlipidemia were more prevalent in hypertensive women compared to men (P < 0.0001).
An earlier age of HTN onset was significantly associated with smoking (P = 0.00007), obesity (BMI ≥ 30 kg/m2; P =
0.0003), increased stress (P = 0.0003) and alcohol consumption (P = 0.004).
Conclusion: Understanding the clinical profile of HTN patients contributes to CVD management. Estonian
hypertension patients exhibited different disease and risk profiles of male and female patients. This wellcharacterized
sample set provides a good resource for studying hypertension and other cardiovascular phenotypes.

PloS one, 2016