Arabidopsis has over 80 genes encoding conserved and plant-specific core cell cycle regulators, but in most cases neither their timing of expression in the cell cycle is known nor whether they ...represent redundant and/or tissue-specific functions. Here we identify novel cell cycle regulators, including new cyclin-dependent kinases related to the mammalian galactosyltransferase-associated protein kinase p58, and new classes of cyclin-like and CDK-like proteins showing strong tissue specificity of expression. We analyse expression of all cell cycle regulators in synchronized Arabidopsis cell cultures using multiple approaches including Affymetrix microarrays, massively parallel signature sequencing and real-time reverse transcriptase polymerase chain reaction, and in plant material using the results of over 320 microarray experiments. These global analyses reveal that most core cell cycle regulators are expressed across almost all tissues and more than 85% are expressed at detectable levels in the cell suspension culture, allowing us to present a unified model of transcriptional regulation of the plant cell cycle. Characteristic patterns of D-cyclin expression in early and late G1 phase, either limited to the re-entry cycle or continuously oscillating, suggest that several CYCD genes with strong oscillatory regulation in late G1 may play the role of cyclin E in plants. Alone amongst the six groups of A and B type cyclins, members of CYCA3 peak in S-phase suggest it is a major component of S-phase kinases, whereas others show a peak in G2/M. 82 genes share this G2/M regulatory pattern, about half being new candidate mitotic genes of previously unknown function.
Aim
Histidine‐containing dipeptides (HCDs) are pleiotropic homeostatic molecules with potent antioxidative and carbonyl quenching properties linked to various inflammatory, metabolic, and ...neurological diseases, as well as exercise performance. However, the distribution and metabolism of HCDs across tissues and species are still unclear.
Methods
Using a sensitive UHPLC–MS/MS approach and an optimized quantification method, we performed a systematic and extensive profiling of HCDs in the mouse, rat, and human body (in n = 26, n = 25, and n = 19 tissues, respectively).
Results
Our data show that tissue HCD levels are uniquely produced by carnosine synthase (CARNS1), an enzyme that was preferentially expressed by fast‐twitch skeletal muscle fibres and brain oligodendrocytes. Cardiac HCD levels are remarkably low compared to other excitable tissues. Carnosine is unstable in human plasma, but is preferentially transported within red blood cells in humans but not rodents. The low abundant carnosine analogue N‐acetylcarnosine is the most stable plasma HCD, and is enriched in human skeletal muscles. Here, N‐acetylcarnosine is continuously secreted into the circulation, which is further induced by acute exercise in a myokine‐like fashion.
Conclusion
Collectively, we provide a novel basis to unravel tissue‐specific, paracrine, and endocrine roles of HCDs in human health and disease.
In darkness, shoot apex growth is repressed, but it becomes rapidly activated by light. We show that phytochromes and cryptochromes play largely redundant roles in this derepression in Arabidopsis ...thaliana. We examined the light activation of transcriptional changes in a finely resolved time course, comparing the shoot apex (meristem and leaf primordia) and the cotyledon and found >5700 differentially expressed genes. Early events specific to the shoot apices included the repression of genes for Really Interesting New Gene finger proteins and basic domain/leucine zipper and basic helix-loop-helix transcription factors. The downregulation of auxin and ethylene and the upregulation of cytokinin and gibberellin hormonal responses were also characteristic of shoot apices. In the apex, genes involved in ribosome biogenesis and protein translation were rapidly and synchronously induced, simultaneously with cell proliferation genes, preceding visible organ growth. Subsequently, the activation of signaling genes and transcriptional signatures of cell wall expansion, turgor generation, and plastid biogenesis were apparent. Furthermore, light regulates the forms and protein levels of two transcription factors with opposing functions in cell proliferation, E2FB and E2FC, through the Constitutively Photomorphogenic1 (COP1), COP9-Signalosome5, and Deetiolated1 light signaling molecules. These data provide the basis for reconstruction of the regulatory networks for light-regulated meristem, leaf, and cotyledon development.
Balenine possesses some of carnosine's and anserine's functions, yet it appears more resistant to the hydrolysing CN1 enzyme. The aim of this study was to elucidate the stability of balenine in the ...systemic circulation and its bioavailability in humans following acute supplementation. Two experiments were conducted in which (in vitro) carnosine, anserine and balenine were added to plasma to compare degradation profiles and (in vivo) three increasing doses (1-4-10 mg/kg) of balenine were acutely administered to 6 human volunteers. Half-life of balenine (34.9 ± 14.6 min) was respectively 29.1 and 16.3 times longer than that of carnosine (1.20 ± 0.36 min, p = 0.0044) and anserine (2.14 ± 0.58 min, p = 0.0044). In vivo, 10 mg/kg of balenine elicited a peak plasma concentration (Cmax) of 28 µM, which was 4 and 18 times higher than with 4 (p = 0.0034) and 1 mg/kg (p = 0.0017), respectively. CN1 activity showed strong negative correlations with half-life (ρ = - 0.829; p = 0.0583), Cmax (r = - 0.938; p = 0.0372) and incremental area under the curve (r = - 0.825; p = 0.0433). Overall, balenine seems more resistant to CN1 hydrolysis resulting in better in vivo bioavailability, yet its degradation remains dependent on enzyme activity. Although a similar functionality as carnosine and anserine remains to be demonstrated, opportunities arise for balenine as nutraceutical or ergogenic aid.
Purpose
Chronic β-alanine supplementation leads to increased levels of muscle histidine-containing dipeptides. However, the majority of ingested β-alanine is, most likely, degraded by two ...transaminases: GABA-T and AGXT2. In contrast to GABA-T, the in vivo role of AGXT2 with respect to β-alanine metabolism is unknown. The purpose of the present work is to investigate if AGXT2 is functionally involved in β-alanine homeostasis.
Methods
Muscle histidine-containing dipeptides levels were determined in AGXT2 overexpressing or knock-out mice and in human subjects with different rs37369 genotypes which is known to affect AGXT2 activity. Further, plasma β-alanine kinetic was measured and urine was obtained from subjects with different rs37369 genotypes following ingestion of 1400 mg β-alanine.
Result
Overexpression of AGXT2 decreased circulating and muscle histidine-containing dipeptides (> 70% decrease;
p
< 0.05), while AGXT2 KO did not result in altered histidine-containing dipeptides levels. In both models, β-alanine remained unaffected in the circulation and in muscle (
p
> 0.05). In humans, the results support the evidence that decreased AGXT2 activity is not associated with altered histidine-containing dipeptides levels (
p
> 0.05). Additionally, following an acute dose of β-alanine, no differences in pharmacokinetic response were measured between subjects with different rs37369 genotypes (
p
> 0.05). Interestingly, urinary β-alanine excretion was 103% higher in subjects associated with lower AGXT2 activity, compared to subjects associated with normal AGXT2 activity (
p
< 0.05).
Conclusion
The data suggest that in vivo, β-alanine is a substrate of AGXT2; however, its importance in the metabolism of β-alanine and histidine-containing dipeptides seems small.
Plant growth is characterised both by continued growth and organogenesis throughout development, as well as by environmental influences on the rate and pattern of these processes. This necessitates a ...close relationship between cell cycle control, differentiation and development that can be readily observed and studied. The sequencing of the Arabidopsis genome has revealed the full complexity of cell cycle regulators in plants, creating a challenge to understand how these genes control plant growth and differentiation, and how they are integrated with intrinsic and external signals. Here, we review the control of the cell cycle and examine how it is integrated with proliferative activity within meristems, and during the differentiation processes leading to leaf and lateral root formation.
The integration of cell division in root growth and development requires mediation of developmental and physiological signals through regulation of cyclin-dependent kinase activity. Cells within the ...pericycle form de novo lateral root meristems, and D-type cyclins (CYCD), as regulators of the G₁-to-S phase cell cycle transition, are anticipated to play a role. Here, we show that the D-type cyclin protein CYCD2;1 is nuclear in Arabidopsis thaliana root cells, with the highest concentration in apical and lateral meristems. Loss of CYCD2;1 has a marginal effect on unstimulated lateral root density, but CYCD2;1 is rate-limiting for the response to low levels of exogenous auxin. However, while CYCD2;1 expression requires sucrose, it does not respond to auxin. The protein Inhibitor-lnteractor of CDK/Kip Related Protein2 (ICK2/KRP2), which interacts with CYCD2;1, inhibits lateral root formation, and ick2/krp2 mutants show increased lateral root density. ICK2/KRP2 can modulate the nuclear levels of CYCD2;1, and since auxin reduces ICK2/KRP2 protein levels, it affects both activity and cellular distribution of CYCD2;1. Hence, as ICK2/KRP2 levels decrease, the increase in lateral root density depends on CYCD2;1, irrespective of ICK2/CYCD2;1 nuclear localization. We propose that ICK2/KRP2 restrains root ramification by maintaining CYCD2;1 inactive and that this modulates pericycle responses to auxin fluctuations.
Introduction:
Acute apnea evokes bradycardia and peripheral vasoconstriction in order to conserve oxygen, which is more pronounced with face immersion. This response is contrary to the tachycardia ...and increased blood flow to muscle tissue related to the higher oxygen consumption during exercise. The aim of this study was to investigate cardiovascular and metabolic responses of dynamic dry apnea (DRA) and face immersed apnea (FIA).
Methods:
Ten female volunteers (17.1 ± 0.6 years old) naive to breath-hold-related sports, performed a series of seven dynamic 30 s breath-holds while cycling at 25% of their peak power output. This was performed in two separate conditions in a randomized order: FIA (15°C) and DRA. Heart rate and muscle tissue oxygenation through near-infrared spectroscopy were continuously measured to determine oxygenated (mO
2
Hb) and deoxygenated hemoglobin concentration (mHHb) and tissue oxygenation index (mTOI). Capillary blood lactate was measured 1 min after the first, third, fifth, and seventh breath-hold.
Results:
Average duration of the seven breath-holds did not differ between conditions (25.3 s ± 1.4 s,
p
= 0.231). The apnea-induced bradycardia was stronger with FIA (from 134 ± 4 to 85 ± 3 bpm) than DRA (from 134 ± 4 to 100 ± 5 bpm,
p
< 0.001). mTOI decreased significantly from 69.9 ± 0.9% to 63.0 ± 1.3% (
p
< 0.001) which is reflected in a steady decrease in mO
2
Hb (
p
< 0.001) and concomitant increase in mHHb (
p
= 0.001). However, this was similar in both conditions (0.121 <
p
< 0.542). Lactate was lower after the first apnea with FIA compared to DRA (
p
= 0.038), while no differences were observed in the other breath-holds.
Conclusion:
Our data show strong decreases in heart rate and muscle tissue oxygenation during dynamic apneas. A stronger bradycardia was observed in FIA, while muscle oxygenation was not different, suggesting that FIA did not influence muscle oxygenation. An order of mechanisms was observed in which, after an initial tachycardia, heart rate starts to decrease after muscle tissue deoxygenation occurs, suggesting a role of peripheral vasoconstriction in the apnea-induced bradycardia. The apnea-induced increase in lactate was lower in FIA during the first apnea, probably caused by the stronger bradycardia.
V0v spinal interneurons are highly conserved, glutamatergic, commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the ...neurotransmitter phenotype of these cells. However, we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells.
To identify candidate members of V0v gene regulatory networks, we FAC-sorted wild-type and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression-profiled them using microarrays and single cell RNA-seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 double mutants and wild-type siblings.
Our data reveal two molecularly distinct subtypes of zebrafish V0v spinal interneurons at 48 h and suggest that, by this stage of development, evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons, or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons, plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly, we show that Hmx2/3a, repress dI2 interneuron expression of skor1a and nefma, two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression.
This study identifies two molecularly distinct subsets of zebrafish V0v spinal interneurons, as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of these cells. Our data further suggest that in the absence of both Evx1 and Evx2, V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then, later, start to express markers of distinct types of inhibitory spinal interneurons, or motoneurons. Taken together, our findings significantly increase our knowledge of V0v and spinal development and move us closer towards the essential goal of identifying the complete gene regulatory networks that specify this crucial cell type.
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