Nucleotide Imbalance, Provoked by Downregulation of Aspartate Transcarbamoylase Impairs Cold Acclimation in Arabidopsis.

Aspartate transcarbamoylase (ATC) catalyzes the first committed step in pyrimidine de novo synthesis. As shown before, mutants with 80% reduced transcript and protein levels exhibit reduced levels of pyrimidine metabolites and thus nucleotide limitation and imbalance. Consequently, reduced photosynthetic capacity and growth, accompanied by massive transcriptional changes, were observed. Here, we show that nucleotide de novo synthesis was upregulated during cold acclimation of Arabidopsis thaliana (ecotype Columbia, Col-0) plants, but ATC knockdown mutants failed to acclimate to this condition as they did not accumulate neutral sugars and anthocyanins. A global transcriptome analysis revealed that most of the transcriptional changes observed in Col-0 plants upon cold exposure were also evident in ATC knockdown plants. However, several responses observed in cold-treated Col-0 plants could already be detected in knockdown plants when grown under standard conditions, suggesting that these mutants exhibited typical cold responses without prior cold stimulation. We believe that nucleotide signaling is involved in "cold-like priming" and "cold acclimation" in general. The observed transcript levels of genes involved in central carbon metabolism and respiration were an exception to these findings. These were upregulated in the cold but downregulated in warm-grown ATC mutants.

Cytidine Triphosphate Synthase Four From Arabidopsis thaliana Attenuates Drought Stress Effects.

Cytidine triphosphate synthase (CTPS) catalyzes the final step in pyrimidine de novo synthesis. In Arabidopsis, this protein family consists of five members (CTPS1-5), and all of them localize to the cytosol. Specifically, CTPS4 showed a massive upregulation of transcript levels during abiotic stress, in line with increased staining of CTPS4 promoter:GUS lines in hypocotyl, root and to lesser extend leaf tissues. In a setup to study progressive drought stress, CTPS4 knockout mutants accumulated less fresh and dry weight at days 5-7 and showed impaired ability to recover from this stress after 3 days of rewatering. Surprisingly, a thorough physiological characterization of corresponding plants only revealed alterations in assimilation and accumulation of soluble sugars including those related to drought stress in the mutant. Bimolecular fluorescence complementation (BiFC) studies indicated the interaction of CTPS4 with other isoforms, possibly affecting cytoophidia (filaments formed by CTPS formation. Although the function of these structures has not been thoroughly investigated in plants, altered enzyme activity and effects on cell structure are reported in other organisms. CTPS activity is required for cell cycle progression and growth. Furthermore, drought can lead to the accumulation of reactive oxygen species (ROS) and by this, to DNA damage. We hypothesize that effects on the cell cycle or DNA repair might be relevant for the observed impaired reduced drought stress tolerance of CTPS4 mutants.

Loss of a pyridoxal-phosphate phosphatase rescues Arabidopsis lacking an endoplasmic reticulum ATP carrier.

The endoplasmic reticulum (ER)-located ATP/ADP-antiporter (ER-ANT1) occurs specifically in vascular plants. Structurally different transporters mediate energy provision to the ER, but the cellular function of ER-ANT1 is still unknown. Arabidopsis (Arabidopsis thaliana) mutants lacking ER-ANT1 (er-ant1 plants) exhibit a photorespiratory phenotype accompanied by high glycine levels and stunted growth, pointing to an inhibition of glycine decarboxylase (GDC). To reveal whether it is possible to suppress this marked phenotype, we exploited the power of a forward genetic screen. Absence of a so far uncharacterized member of the HaloAcid Dehalogenase (HAD)-like hydrolase family strongly suppressed the dwarf phenotype of er-ant1 plants. Localization studies suggested that the corresponding protein locates to chloroplasts, and activity assays showed that the enzyme dephosphorylates, with high substrate affinity, the B6 vitamer pyridoxal 5'-phosphate (PLP). Additional physiological experiments identified imbalances in vitamin B6 homeostasis in er-ant1 mutants. Our data suggest that impaired chloroplast metabolism, but not decreased GDC activity, causes the er-ant1 mutant dwarf phenotype. We present a hypothesis, setting transport of PLP by ER-ANT1 and chloroplastic PLP dephosphorylation in the cellular context. With the identification of this HAD-type PLP phosphatase, we also provide insight into B6 vitamer homeostasis.

Occurrence of Rare Deletional Yus and Gerbich Alleles in Syria and Neighbouring Countries.

Background: Gerbich-negative phenotypes of the Gerbich Blood Group System (ISBT 020) are very rare (with the exception of Papua New Guinea). The Gerbich-negative phenotypes Yus and Gerbich are negative for the antigens Ge2, and Ge2 and Ge3, respectively. In antigen-negative individuals, anti-Ge2 and anti-Ge3 antibodies can be naturally occurring, or are triggered during pregnancies and after transfusions. Previous studies suggested an elevated frequency of Gerbich-negative phenotypes for the Middle East. In the summer of 2015, a large-scale migration of people from the Middle East to Europe occurred raising the issue of question how to guarantee blood supply for patients and manage antenatal care for pregnant women from these countries. Materials and Methods: To investigate the frequency of rare Gerbich-negative phenotypes, 1,665 immigrants to Germany originating from the Middle East were genetically tested for the presence of rare Yus, i.e., GE*01.-02, and Gerbich, i.e., GE*01-03, alleles and compared to results obtained from 507 Germans. Results: Seven Yus GE*01.-02.01 and one Gerbich GE*01.-03.02 alleles were exclusively observed among people from the Middle East, with five of them clustering among 797 Syrians. No such alleles were observed in Germans. A cumulative Yus- and GE*01.-03-type allele frequency of 0.00314 and resultant overall Gerbich-negative phenotype frequency of one among 101,633 Syrians were calculated. Conclusion: This manuscript describes for the first time an exclusively genetic screening for carriers of Gerbich-negative alleles. In conclusion, the Gerbich blood group system should be considered as one causative agent of unusual antibodies to red cell antigens, in routine patients and pregnant women, especially when originating from the Middle East.

Cytosolic CTP Production Limits the Establishment of Photosynthesis in Arabidopsis.

CTP synthases (CTPS) comprise a protein family of the five members CTPS1-CTPS5 in Arabidopsis, all located in the cytosol. Specifically, downregulation of CTPS2 by amiRNA technology results in plants with defects in chlorophyll accumulation and photosynthetic performance early in development. CTP and its deoxy form dCTP are present at low levels in developing seedlings. Thus, under conditions of fast proliferation, the synthesis of CTP (dCTP) can become a limiting factor for RNA and DNA synthesis. The higher sensitivity of ami-CTPS2 lines toward the DNA-Gyrase inhibitor ciprofloxacin, together with reduced plastid DNA copy number and 16S and 23S chloroplast ribosomal RNA support this view. High expression and proposed beneficial biochemical features render CTPS2 the most important isoform for early seedling development. In addition, CTPS2 was identified as an essential enzyme in embryo development before, as knock-out mutants were embryo lethal. In line with this, ami-CTPS2 lines also exhibited reduced seed numbers per plant.

Characterization of the novel HLA-DPB1*1219:01 allele by next-generation sequencing.

The new allele differs from DPB1*296:01 by a c.292G>A substitution in exon 2.

Platelet CD36 deficiency is present in 2.6% of Arabian individuals and can cause NAIT and platelet refractoriness.

BACKGROUND: CD36 isoantibodies are capable of inducing neonatal alloimmune thrombocytopenia (NAIT) and platelet refractoriness. As to now the CD36 type I deficiency has been reported in East Asian and African individuals. However, it is virtually unknown in Caucasians. The aim of this study was to display the prevalence of the CD36 deficiency within parts of the Arabian population in Germany. Secondly, we are presenting the case of a newborn suffering from NAIT which was induced by CD36 antibody. METHODS: Platelet (p) CD36 was determined by flow cytometry on 1328 samples mainly from individuals of Arabian origin and a family with a neonate affected by NAIT. DNA sequencing was performed on all pCD36-negative samples. RESULTS: Thirty-five (2.64%) of all donor samples were pCD36 negative, 19 (1.43%) had a weak expression. Including only individuals from the Arabian peninsula, frequencies were 3.39% and 1.75%, respectively. CD36 type I deficiency on both platelets and monocytes combined with a CD36 isoantibody were detected in the mother of the NAIT baby. The baby was successfully transfused with two HPA-unselected platelet concentrates. In case of need, two platelet units with a weak pCD36 expression were on hand. A total of 45 different CD36 mutations were detected within pCD36-negative individuals, some being homozygous, most of them only present on one allele. CONCLUSION: The CD36-negative phenotype is present in a significant number of individuals of Arabian origin and enables CD36 isoimmunization in NAIT or refractoriness. Blood transfusion services should be aware of such cases.

Molecular Blood Group Screening in Donors from Arabian Countries and Iran Using High-Throughput MALDI-TOF Mass Spectrometry and PCR-SSP.

BACKGROUND AND AIMS: Only little is known about blood groups other than ABO blood groups and Rhesus factors in Arabian countries and Iran. During the last years, increased migration to Central Europe has put a focus on the question how to guarantee blood supply for patients from these countries, particularly because hemoglobinopathies with the need of regular blood support are more frequent in patients from that region. Therefore, blood group allele frequencies should be determined in individuals from Arabian countries and Iran by molecular typing and compared to a German rare donor panel. METHODS: 1,111 samples including 800 individuals from Syria, 147 from Iran, 123 from the Arabian Peninsula, and 41 from Northern African countries were included in a MALDI-TOF MS assay to detect polymorphisms coding for Kk, Fy(a/b), Fynull, Cw, Jk(a/b), Jo(a+/a-), Lu(a/b), Lu(8/14), Ss, Do(a/b), Co(a/b), In(a/b), Js(a/b), Kp(a/b), and variant alleles RHCE*c.697C>G and RHCE *c.733C>G. Yt(a/b), S-s-U-, Velnull, Conull, and RHCE *c.667G>T were tested by PCR-SSP. RESULTS: Of the Arabian donors, 2% were homozygous for the FY *02.01N allele (Fynull), and 15.7% carried the heterozygous mutation. However, 0.8% of the German donors also carried 1 copy of the allele. 3.6% of all and 29.3% of Northern African donors were heterozygous for the RHCE *c.733C>G substitution, 0.4% of the Syrian probands were heterozygous for DO *01/DO *01.-05, a genotype that was lacking in German donors. Whereas the KEL *02.06 allele coding for the Js(a) phenotype was missing in Germans; 0.8% of the Syrian donors carried 1 copy of this allele. 1.8% of the Syrian but only 0.3% of the German donors were negative for YT *01. One donor from Northern Africa homo-zygously carried the GYPB *270+5g>t mutation, inducing the S-s-U+w phenotype, and in 2 German donors a GYPB *c.161G>A exchange, which induces the Mit+ phenotype, caused a GYPB *03 allele dropout in the MALDI assay. The overall failure rate of the Arabian panel was 0.4%. CONCLUSIONS: Some blood group alleles that are largely lacking in Europeans but had been described in African individuals are present in Arabian populations at a somewhat lower frequency. In single cases, it could be challenging to provide immunized Arabian patients with compatible blood.

Endoscopic Ultrasound Features of Multiple Endocrine Neoplasia Type 1-Related versus Sporadic Pancreatic Neuroendocrine Tumors: A Single-Center Retrospective Study.

AIM: Pancreatic neuroendocrine tumors (pNETs) can occur in patients with a familial syndrome either as multiple endocrine neoplasia type 1 (MEN-1) or as sporadic tumors. Endoscopic ultrasound (EUS) has become one of the first-line investigations for pNET characterization. The ultrasonographic features of pNETs may differ depending on the familial versus sporadic pathogenesis of the tumor. Therefore, the EUS findings could help and direct the definition of a pNET with an impact on the most appropriate diagnostic and therapeutic patient management. METHODS: In this single-center retrospective study, we reviewed the EUS features of 94 pNETs from 37 MEN-1 patients and 15 pNETs from 11 sporadic disease patients at the time of their first EUS assessment. We analyzed the most relevant morphological and ultrasonographic characteristics of the tumors and compared the findings between the 2 patient groups. RESULTS: Patients with MEN-1 more likely present with multiple pNETs than patients with sporadic disease. Sporadic pNETs are usually much bigger than those due to MEN-1. Moreover, pNETs are more heterogeneous in patients with sporadic disease than in those with MEN-1. No statistical difference with regard to definition of the margins, morphology, and vascularization of the pNETs appears between the 2 groups. CONCLUSIONS: Patients with sporadic disease usually present with bigger and more heterogeneous pNETs than patients with MEN-1, who tend to present with a higher number of lesions. EUS can facilitate the precise characterization of a pNET, and the ultrasonographic features of the lesion can help and distinguish MEN-1-related versus sporadic disease.