Apple (Malus × domestica) trees are vulnerable to freezing temperatures. However, there has been only limited success in developing cold-hardy cultivars. This lack of progress is due at least partly ...to lack of understanding of the molecular mechanisms of freezing tolerance in apple.
In this study, we evaluated the potential roles for two R2R3 MYB transcription factors (TFs), MYB88 and the paralogous FLP (MYB124), in cold stress in apple and Arabidopsis. We found that MYB88 and MYB124 positively regulate freezing tolerance and cold-responsive gene expression in both apple and Arabidopsis.
Chromatin-Immunoprecipitation-qPCR and electrophoretic mobility shift assays showed that MdMYB88/MdMYB124 act as direct regulators of the COLD SHOCK DOMAIN PROTEIN 3 (MdCSP3) and CIRCADIAN CLOCK ASSOCIATED 1 (MdCCA1) genes. Dual luciferase reporter assay indicated that MdCCA1 but not MdCSP3 activated the expression of MdCBF3 under cold stress. Moreover, MdMYB88 and MdMYB124 promoted anthocyanin accumulation and H2O2 detoxification in response to cold.
Taken together, our results suggest that MdMYB88 and MdMYB124 positively regulate cold hardiness and cold-responsive gene expression under cold stress by C-REPEAT BINDING FACTOR (CBF)-dependent and CBF-independent pathways.
Abstract
MdMYB88 and MdMYB124 have been demonstrated to be responsible for lignin accumulation in apple under drought stress. In this study, using a metabolomic approach, we identified differentially ...accumulated phenylpropanoid and flavonoid metabolites in
MdMYB88/124
transgenic RNAi plants under control and long-term drought stress conditions in apple roots. We confirmed the regulation of phenylalanine by MdMYB88 and MdMYB124 via UPLC-MS in apple roots under both control and drought conditions. Using Electrophoretic Mobility Shift Assay (EMSA) and ChIP-quantitative PCR (qPCR) analyses, we found that MdMYB88 positively regulates the MdCM2 gene, which is responsible for phenylalanine biosynthesis, through binding to its promoter region. Under long-term drought conditions,
MdMYB88/124
RNAi plants consistently accumulated increased amounts of H
2
O
2
and MDA, while
MdMYB88
and
MdMYB124
overexpression plants accumulated decreased amounts of H
2
O
2
and MDA. We also examined the accumulation of metabolites in the phenylpropanoid biosynthesis pathway in the leaves of
MdMYB88
and
MdMYB124
transgenic apple plants after long-term drought stress. We found that metabolites responsible for plant defense, including phenylpropanoids and flavonoids, accumulated less in the RNAi plants but more in the overexpression plants under both control and drought conditions. We further demonstrated that
MdMYB88/124
RNAi plants were more sensitive to
Alternaria alternata
f. sp.
mali
and
Valsa mali
, two pathogens that currently severely threaten apple production. In contrast,
MdMYB88
and
MdMYB124
overexpression plants were more tolerant to these pathogens. The cumulative results of this study provided evidence for secondary metabolite regulation by MdMYB88 and MdMYB124, further explained the molecular roles of MdMYB88 and MdMYB124 in drought resistance, and provided information concerning molecular aspects of their roles in disease resistance.
Drought stress can negatively impact apple fruit quality and yield. Apple microRNAs (miRNAs) participate in apple tree and fruit development, as well as in biotic stress tolerance; however, it is ...largely unknown whether these molecules are involved in the drought response. To identify drought-responsive miRNAs in
, we first examined the drought stress tolerance of ten F
progenies of R3 (
. ×
) ×
. We performed Illumina sequencing on pooled total RNA from both drought-tolerant and drought-sensitive plants. The sequencing results identified a total of 206 known miRNAs and 253 candidate novel miRNAs from drought-tolerant plants and drought-sensitive plants under control or drought conditions. We identified 67 miRNAs that were differentially expressed in drought-tolerant plants compared with drought-sensitive plants under drought conditions. Under drought stress, 61 and 35 miRNAs were differentially expressed in drought-tolerant and drought-sensitive plants, respectively. We determined the expression levels of seven out of eight miRNAs by stem-loop qPCR analysis. We also predicted the target genes of all differentially expressed miRNAs and identified the expression of some genes. Gene Ontology analyses indicated that the target genes were mainly involved in stimulus response and cellular and metabolic processes. Finally, we confirmed roles of two miRNAs in apple response to mannitol. Our results reveal candidate miRNAs and their associated mRNAs that could be targeted for improving drought tolerance in
species, thus providing a foundation for understanding the molecular networks involved in the response of apple trees to drought stress.
Noonan syndrome (NS) is a relatively rare inherited disease. Typical clinical presentation is important for the diagnosis of NS. But the initial presentation of NS could be significant variant ...individually which results in the difficult of working diagnosis. Here we report a rare neonatal case of NS who presented with refractory thrombocytopenia as the initial manifestation.
This was a preterm infant with refractory thrombocytopenia of unknown origin transferred from obstetric hospital at 6 weeks of age. During hospitalization, typical phenotypes of NS in addition to thrombocytopenia were observed, such as typical facial characteristics, short stature, atrial septal defect, cryptochidism, coagulation defect and chylothorax. Genetic testing showed a pathogenic variant at exon 2 of the PTPN11 gene with c.124A > G (p.T42A). Respiratory distress was deteriorated with progressive chylothorax. Chest tube was inserted for continuous draining. Chemical pleurodesis with erythromycin was tried twice, but barely effective. Finally, parents decided to withdraw medical care and the patient died.
Thrombocytopenia could be the first symptom of Noonan syndrome. After ruling out other common causes of thrombocytopenia, NS should be considered as the working diagnosis.
To investigate if insulin resistance per se or the accompanying hyperinsulinemia induced hypertension and its underlying mechanisms.
Sprague-Dawley rats were randomized into normal diet-fed group (ND ...group) and high-fat diet-fed group (HFD group). Then, the HFD group was further randomly divided into the control group (HFD_C group), the PIO group (treated with pioglitazone), the STZ_DM group (to induce diabetes with streptozotocin) and the DM+Ins group (streptozotocin injection followed by insulin treatment). Insulin sensitivity, plasma insulin, endothelin-1, norepinephrine, aldosterone, angiotensinⅡ and 24-h urinary sodium excretion (USE) levels of the groups were measured and analyzed. A multiple stepwise regression analysis method was applied to exam our hypothesis.
Compared to HFD_C group, the groups with lower plasma insulin, the PIO group and STZ_DM group, showed higher USE and lower blood pressure. The groups with higher plasma insulin (but same level of insulin resistance), the HFD_C group and DM+Ins group, showed lower USE and higher blood pressure. The 24-h urinary sodium excretion was the most important contributor to the significant changes of blood pressure with an R
2
of 25.2% in this animal experiment.
It is the compensatory hyperinsulinemia rather than insulin resistance per se that causes blood pressure elevation. The urinary sodium excretion is the key mediator among the multiple mechanisms. Therapies targeting hyperinsulinemia and restricting salt intake may favor a better control of hypertension associated with insulin resistance.
Nucleic acids in plant tissue lysates can be captured quickly by a cellulose filter paper and prepared for amplification after a quick purification. In this study, a published filter paper strip ...method was modified by sticking the filter paper on a polyvinyl chloride resin (PVC) sheet. This modified method is named EZ-D, for EASY DNA extraction. Compared with the original cetyl trimethylammonium bromide (CTAB) method, DNA extracted by EZ-D is more efficient in polymerase chain reaction (PCR) amplification due to the more stable performance of the EZ-D stick. The EZ-D method is also faster, easier, and cheaper. PCR analyses showed that DNA extracted from several types of plant tissues by EZ-D was appropriate for specific identification of biological samples. A regular PCR reaction can detect the EZ-D-extracted DNA template at concentration as low as 0.1 ng/µL. Evaluation of the EZ-D showed that DNA extracts could be successfully amplified by PCR reaction for DNA fragments up to 3000 bp in length and up to 80% in GC content. EZ-D was successfully used for DNA extraction from a variety of plant species and plant tissues. Moreover, when EZ-D was combined with the loop-mediated isothermal amplification (LAMP) method, DNA identification of biological samples could be achieved without the need for specialized equipment. As an optimized DNA purification method, EZ-D shows great advantages in application and can be used widely in laboratories where equipment is limited and rapid results are required.
This study aimed to assess whether a higher insulin response increased the long-term risk of mortality in a non-diabetic population.
A total of 446 people with normal glucose tolerance (NGT) or ...impaired glucose tolerance (IGT) who participated in the Da Qing Diabetes Study, were stratified into quartiles subgroups according to their baseline insulin area under the curve (AUC) during oral glucose tolerance test, defined as Q1, Q2, Q3 and Q4. The participants were followed from 1986 to 2016 to assess the risk of death in association with the magnitude of post-load insulin response.
Over 30 years, the rates of all cause death were 9.94, 14.81, 15.02, and 17.58 per 1000 person-years across the four groups respectively. The rate for cardiovascular disease (CVD) death was 5.14, 6.50, 6.80 and 10.47 per 1000 person-years. Compared with Q1, the risk of all-cause death was significantly higher in participants in Q4 (HR = 2.14, 95% CI 1.34-3.42), Q3 (HR = 1.94, 95% CI 1.20-3.14), and Q2 group (HR = 1.70, 95% CI 1.06-2.74). In the Fine-Gray model with non-CVD death as competing risk, the increased insulin AUC were also significantly associated with the CVD death (Q4 vs Q1, HR = 2.04, 95% CI 1.10-3.79). In the fractional polynomial regression analysis, a nonlinear association between insulin AUC and all-cause and CVD death was demonstrated. In addition, insulin AUC was associated with a progressively higher risk of all-cause death and CVD death (fractional power 3, P < 0.001).
A higher post-load insulin response was significantly associated with a long-term increased risk of all-cause and CVD deaths in the Chinese non-diabetic population. It suggests that people featured by this phenotype is a potential important target for further intervention.
Nitrilase is a valuable type of hydrolase that catalyzes nitriles into carboxylic acid and ammonia. Its applications, however, are severely restricted by the harsh conditions of industrial reaction ...processes. To solve this problem, a nitrilase from
72W was inserted into an
shuttle vector for spore surface display. Western blot, enzyme activity measurements and flow cytometric analysis results all indicated a successful spore surface display of the CotB-nit fusion protein. In addition, the optimal catalytic pH value and temperature of the displayed nitrilase were determined to be 7.0 and 50°C, respectively. Moreover, results of reusability tests revealed that 64% of the initial activity of the displayed nitrilase was still retained at the 10
cycle. Furthermore, hydrolysis efficiency of upscale production of cyanocarboxylic acid was significantly higher in the displayed nitrilase-treated group than in the free group expressed by
(pET-28a-nit). Generally, the display of
72W nitrilase on the spore surface of
may be a useful method for immobilization of enzyme and consequent biocatalytic stabilization.
Persimmon anthracnose, a severe disease caused by the hemibiotrophic fungus
Colletotrichum horii
, poses a substantial threat to China’s persimmon industry. Previous research showed that ‘Kangbing ...Jianshi’ cultivar exhibits strong resistance to anthracnose. Notably, ‘Kangbing Jianshi’ branches exhibit greater lignification compared with the susceptible ‘Fuping Jianshi’ cultivar. In this study, higher lignin content was observed in ‘Kangbing Jianshi’ compared with ‘Fuping Jianshi’, and this difference was associated with disease resistance. Transcriptome and metabolome analyses revealed that the majority of differentially expressed genes and differentially accumulated metabolites were primarily enriched in the phenylpropanoid biosynthesis and lignin synthesis pathways. Furthermore, significant upregulation of
DkCAD1
, a pivotal gene involved in lignin metabolism, was observed in the resistant cultivar when inoculated with
C. horii
. Transient overexpression of
DkCAD1
substantially increased lignin content and improved resistance to
C. horii
in a susceptible cultivar. Furthermore, through yeast one-hybrid (Y1H) assays, we identified two WRKY transcription factors, DkWRKY8 and DkWRKY10, which interacts with the
DkCAD1
promoter and induces its activity. Overexpression of
DkWRKY8
and
DkWRKY10
not only increased leaf lignin content but also enhanced persimmon tolerance to
C
.
horii
. Moreover, the expression levels of
DkCAD1
,
DkWRKY8
, and
DkWRKY10
were significantly increased in response to salicylic acid and jasmonic acid in the resistant cultivar. These findings enhance our understanding of the molecular functions of
DkWRKY8
,
DkWRKY10
, and
DkCAD1
in persimmons, as well as their involvement in molecular breeding processes in persimmons.
Background
Limited information is available on the long‐term risk of cardiovascular disease (CVD) associated with hypertriglyceridaemia (HTG) in the Chinese population. We estimated this risk over a ...23‐year period in participants recruited from among those included in the Da Qing Diabetes Study.
Methods
A total of 833 Chinese adults including 379 with normal glucose levels and 454 with hyperglycaemia were identified by their oral glucose tolerance in 1986 in Da Qing, China. CVD outcomes were monitored until 2009. Thirty‐four percent (280/833) of the participants had HTG, which was defined as a fasting plasma triglyceride (TG) level ≥ 1.7 mmol/L, at the baseline time point.
Results
Over the 23‐yearfollow‐up period, 149 subjects in the HTG group and 190 subjects in the non‐HTG group (NTG group) experienced their first CVD event, including fatal or nonfatal myocardial infarction (MI) and stroke. The age and sex‐adjusted annual incidence of the first CVD event per 1000 person‐years was 30.23 for the HTG group vs 18.68 for the NTG group. The corresponding rates for MI and stroke were 7.71 vs 3.89 and 19.55 vs 13.98, respectively. After adjusting for confounders, the HTG group had a 28% higher risk of the first CVD event than the NTG group. This association was significant among only the subjects with a serum cholesterol level > 5.7 mmol/L and those with diabetes or impaired glucose tolerance (IGT).
Conclusion
HGT predicted a substantially higher subsequent long‐term risk of the first CVD event in Chinese adults, especially in those with hypercholesterolaemia and hyperglycaemia.
PDF
