Dyrk1A, a serine/threonine kinase, is involved in ERK and Akt activation in the brain of hyperhomocysteinemic mice.

Hyperhomocysteinemia due to cystathionine beta synthase (CBS) deficiency is associated with diverse brain disease. Whereas the biological actions linking hyperhomocysteinemia to the cognitive dysfunction are not well understood, we tried to establish relationships between hyperhomocysteinemia and alterations of signaling pathways. In the brain of CBS-deficient mice, a murine model of hyperhomocysteinemia, we previously found an activation of extracellular signal-regulated kinase (ERK) pathway and an increase of Dyrk1A, a serine/threonine kinase involved in diverse functions ranging from development and growth to apoptosis. We then investigated the relationship between Dyrk1A and the signaling pathways initiated by receptor tyrosine kinases (RTK), the ERK and PI3K/Akt pathways. We found a significant increase of phospho-ERK, phospho-MEK, and phospho-Akt in the brain of CBS-deficient and Dyrk1a-overexpressing mice. This increase was abolished when CBS-deficient and Dyrk1A-transgenic mice were treated with harmine, an inhibitor of Dyrk1A kinase activity, which emphasizes the role of Dyrk1A activity on ERK and Akt activation. Sprouty 2 protein level, a negative feedback loop modulator that limits the intensity and duration of RTK activation, is decreased in the brain of CBS-deficient mice, but not in the brain of Dyrk1A transgenic mice. Furthermore, a reduced Dyrk1A and Grb2 binding on sprouty 2 and an increased interaction of Dyrk1A with Grb2 were found in the brain of Dyrk1A transgenic mice. The consequence of Dyrk1A overexpression on RTK activation seems to be a decreased interaction of sprouty 2/Grb2. These observations demonstrate ERK and Akt activation induced by Dyrk1A in the brain of hyperhomocysteinemic mice and open new perspectives to understand the basis of the cognitive defects in hyperhomocysteinemia.

Mice deficient in cystathionine beta synthase display increased Dyrk1A and SAHH activities in brain.

Hyperhomocysteinemia is associated with brain disease. However, biological actions linking hyperhomocysteinemia to neuronal abnormalities are not well understood. We recently found a relationship between Dyrk1A protein expression, a serine/threonine kinase that might be responsible for cognitive functions in Down's syndrome, and hepatic S-adenosylhomocysteine hydrolase (SAHH) activity, which plays a key role in S-adenosylmethionine-dependent methylation reactions. Considering the role of methylation and Dyrk1A in cognitive functions, the aim of this study was to investigate the relationship between Dyrk1A and SAHH activity in brain of hyperhomocysteinemic mice. We found an increase in Dyrk1A protein expression and activity in brain of hyperhomocysteinemic mice, concomitant with an increased SAHH activity. The effect of overexpression of protein Dyrk1A on SAHH activity was confirmed in brain of Dyrk1A transgenic mice, and additionally we found a positive correlation between Dyrk1A and SAHH activity. These observations suggest a potential effect of Dyrk1A on brain phenotypes linked to hyperhomocysteinemia.

Molecular signatures of cardiac defects in Down syndrome lymphoblastoid cell lines suggest altered ciliome and Hedgehog pathways.

Forty percent of people with Down syndrome exhibit heart defects, most often an atrioventricular septal defect (AVSD) and less frequently a ventricular septal defect (VSD) or atrial septal defect (ASD). Lymphoblastoid cell lines (LCLs) were established from lymphocytes of individuals with trisomy 21, the chromosomal abnormality causing Down syndrome. Gene expression profiles generated from DNA microarrays of LCLs from individuals without heart defects (CHD(-); n = 22) were compared with those of LCLs from patients with cardiac malformations (CHD(+); n = 21). After quantile normalization, principal component analysis revealed that AVSD carriers could be distinguished from a combined group of ASD or VSD (ASD+VSD) carriers. From 9,758 expressed genes, we identified 889 and 1,016 genes differentially expressed between CHD(-) and AVSD and CHD(-) and ASD+VSD, respectively, with only 119 genes in common. A specific chromosomal enrichment was found in each group of affected genes. Among the differentially expressed genes, more than 65% are expressed in human or mouse fetal heart tissues (GEO dataset). Additional LCLs from new groups of AVSD and ASD+VSD patients were analyzed by quantitative PCR; observed expression ratios were similar to microarray results. Analysis of GO categories revealed enrichment of genes from pathways regulating clathrin-mediated endocytosis in patients with AVSD and of genes involved in semaphorin-plexin-driven cardiogenesis and the formation of cytoplasmic microtubules in patients with ASD-VSD. A pathway-oriented search revealed enrichment in the ciliome for both groups and a specific enrichment in Hedgehog and Jak-stat pathways among ASD+VSD patients. These genes or related pathways are therefore potentially involved in normal cardiogenesis as well as in cardiac malformations observed in individuals with trisomy 21.

BDNF and DYRK1A are variable and inversely correlated in lymphoblastoid cell lines from Down syndrome patients.

Down syndrome or trisomy 21 is the most common genetic disorder leading to mental retardation. One feature is impaired short- and long-term spatial memory, which has been linked to altered brain-derived neurotrophic factor (BDNF) levels. Mouse models of Down syndrome have been used to assess neurotrophin levels, and reduced BDNF has been demonstrated in brains of adult transgenic mice overexpressing Dyrk1a, a candidate gene for Down syndrome phenotypes. Given the link between DYRK1A overexpression and BDNF reduction in mice, we sought to assess a similar association in humans with Down syndrome. To determine the effect of DYRK1A overexpression on BDNF in the genomic context of both complete trisomy 21 and partial trisomy 21, we used lymphoblastoid cell lines from patients with complete aneuploidy of human chromosome 21 (three copies of DYRK1A) and from patients with partial aneuploidy having either two or three copies of DYRK1A. Decreased BDNF levels were found in lymphoblastoid cell lines from individuals with complete aneuploidy as well as those with partial aneuploidies conferring three DYRK1A alleles. In contrast, lymphoblastoid cell lines from individuals with partial trisomy 21 having only two DYRK1A copies displayed increased BDNF levels. A negative correlation was also detected between BDNF and DYRK1A levels in lymphoblastoid cell lines with complete aneuploidy of human chromosome 21. This finding indicates an upward regulatory role of DYRK1A expression on BDNF levels in lymphoblastoid cell lines and emphasizes the role of genetic variants associated with psychiatric disorders.

Trisomy for synaptojanin1 in Down syndrome is functionally linked to the enlargement of early endosomes.

Enlarged early endosomes have been observed in neurons and fibroblasts in Down syndrome (DS). These endosome abnormalities have been implicated in the early development of Alzheimer's disease (AD) pathology in these subjects. Here, we show the presence of enlarged endosomes in blood mononuclear cells and lymphoblastoid cell lines (LCLs) from individuals with DS using immunofluorescence and confocal microscopy. Genotype-phenotype correlations in LCLs carrying partial trisomies 21 revealed that triplication of a 2.56 Mb locus in 21q22.11 is associated with the endosomal abnormalities. This locus contains the gene encoding the phosphoinositide phosphatase synaptojanin 1 (SYNJ1), a key regulator of the signalling phospholipid phosphatidylinositol-4,5-biphosphate that has been shown to regulate clathrin-mediated endocytosis. We found that SYNJ1 transcripts are increased in LCLs from individuals with DS and that overexpression of SYNJ1 in a neuroblastoma cell line as well as in transgenic mice leads to enlarged endosomes. Moreover, the proportion of enlarged endosomes in fibroblasts from an individual with DS was reduced after silencing SYNJ1 expression with RNA interference. In LCLs carrying amyloid precursor protein (APP) microduplications causing autosomal dominant early-onset AD, enlarged endosomes were absent, suggesting that APP overexpression alone is not involved in the modification of early endosomes in this cell type. These findings provide new insights into the contribution of SYNJ1 overexpression to the endosomal changes observed in DS and suggest an attractive new target for rescuing endocytic dysfunction and lipid metabolism in DS and in AD.

Dyrk1a activates antioxidant NQO1 expression through an ERK1/2-Nrf2 dependent mechanism.

BACKGROUND AND AIMS: Among cardiovascular risk factor, people with Down syndrome have a lower plasma homocysteine level. In a previous study, we have shown that DYRK1A (dual-specificity tyrosine-(Y)-phosphorylation regulated kinase 1a), a serine/threonine kinase found on human chromosome 21, is implicated on homocysteine metabolism regulation. Indeed, mice that overexpress in liver this kinase have a lower plasma homocysteine level concomitant with an increased hepatic S-adenosyhomocysteine hydrolase (SAHH) activity, which depends on the activation of NAD(P)H:quinone oxidoreductase-1 (NQO1). Since NQO1 gene transcription is under the control of NRF2 and AhR, the aim of the present study was to analyze the effect of DYRK1A overexpression in mice onto NRF2 and AhR signaling pathways. METHODS: Effects of DYRK1A overexpression were examined in mice overexpressing Dyrk1a treated with an inhibitor, harmine, by real-time quantitative reverse-transcription polymerase reaction and western blotting. RESULTS: We found that overexpression of DYRK1A increases the nuclear NRF2 quantity, concomitant with the activation of ERK1/2. We also show that the overexpression of Dyrk1a has no effect on PI3K/AKT activation, and AhR signaling pathway in liver of mice. CONCLUSIONS: Our results reveal a link between DYRK1A and NRF2 signaling pathway.

Effect of red wine polyphenol dietary supplementation on two phase II enzymes in liver of hyperhomocysteinemic mice.

Hyperhomocysteinemia leads to diverse clinical manifestations, notably liver disease. The pathogenicity of homocysteine is believed to be due to its ability to produce oxidative stress. Paraoxonase-1 (Pon1), a phase I xenobiotic-metabolizing enzyme (XME) synthesized by liver with anti-oxidative properties within the circulating system is down regulated in case of hyperhomocysteinemia. In a previous study, we have shown that red wine polyphenol extract (PE) supplementation induces a decrease in plasma homocysteine level and an increase in hepatic Pon1 gene expression concomitant with an increase in hepatic and plasma Pon1 activity in a murine model of hyperhomocysteinemia. In the present study, we analyzed the effect of PE supplementation on two phase II XME: NAD(P)H:quinone oxidoreductase (Nqo1) and arylamine-N-acetyltransferase (Nat) family. We found that hyperhomocysteinemia leads to a decrease of hepatic Nqo1 gene expression and activity with a reversal effect of PE supplementation. We also found that hyperhomocysteinemia-induced decrease of peroxynitrite level is associated with an increase of hepatic total Nat activity mainly due to the Nat2 isoform with a reversal effect of PE supplementation. Our results show a beneficial effect of PE supplementation on two phase II enzymes which are altered in case of hyperhomocysteinemia.

DYRK1A, a novel determinant of the methionine-homocysteine cycle in different mouse models overexpressing this Down-syndrome-associated kinase.

BACKGROUND: Hyperhomocysteinemia, characterized by increased plasma homocysteine level, is associated with an increased risk of atherosclerosis. On the contrary, patients with Down syndrome appear to be protected from the development of atherosclerosis. We previously found a deleterious effect of hyperhomocysteinemia on expression of DYRK1A, a Down-syndrome-associated kinase. As increased expression of DYRK1A and low plasma homocysteine level have been associated with Down syndrome, we aimed to analyze the effect of its over-expression on homocysteine metabolism in mice. METHODOLOGY/PRINCIPAL FINDINGS: Effects of DYRK1A over-expression were examined by biochemical analysis of methionine metabolites, real-time quantitative reverse-transcription polymerase chain reaction, and enzyme activities. We found that over-expression of Dyrk1a increased the hepatic NAD(P)H:quinone oxidoreductase and S-adenosylhomocysteine hydrolase activities, concomitant with decreased level of plasma homocysteine in three mice models overexpressing Dyrk1a. Moreover, these effects were abolished by treatment with harmine, the most potent and specific inhibitor of Dyrk1a. The increased NAD(P)H:quinone oxidoreductase and S-adenosylhomocysteine hydrolase activities were also found in lymphoblastoid cell lines from patients with Down syndrome. CONCLUSIONS/SIGNIFICANCE: Our results might give clues to understand the protective effect of Down syndrome against vascular defect through a decrease of homocysteine level by DYRK1A over-expression. They reveal a link between the Dyrk1a signaling pathway and the homocysteine cycle.

Effect of hyperhomocysteinemia on the protein kinase DYRK1A in liver of mice.

Hyperhomocysteinemia due to cystathionine beta synthase (CBS)-deficiency confers diverse clinical manifestations, notably liver diseases. Even if hyperhomocysteinemia in liver of CBS-deficient mice, a murine model of hyperhomocysteinemia, promotes mitochondrial oxidative stress and pro-apoptotic signals, protective signals may counteract these pro-apoptotic signals, leading to chronic inflammation. As DYRK1A, a serine/threonine kinase, has been described as a candidate antiapoptotic factor, we have analyzed the expression of DYRK1A in liver of CBS-deficient mice. We found that DYRK1A protein level was reduced in liver of CBS-deficient mice, which was not observed at the gene expression level. Moreover, the use of primary hepatocytes/Kupffer cells co-culture showed that degradation of DYRK1A induced by hyperhomocysteinemia requires calpain activation. Our results demonstrate a deleterious effect of hyperhomocysteinemia on DYRK1A protein expression, and emphasize the role of hyperhomocysteinemia on calpain activation.