Poly(C)-Binding Protein Pcbp2 Enables Differentiation of Definitive Erythropoiesis by Directing Functional Splicing of the Runx1 Transcript

ABSTRACTFormation of the mammalian hematopoietic system is under a complex set of developmental controls. Here, we report that mouse embryos lacking the KH domain poly(C) binding protein, Pcbp2, are selectively deficient in the definitive erythroid lineage. Compared to wild-type controls, transcript splicing analysis of the Pcbp2−/−embryonic liver reveals accentuated exclusion of an exon (exon 6) that encodes a highly conserved transcriptional control segment of the hematopoietic master regulator, Runx1. Embryos rendered homozygous for a Runx1 locus lacking this cassette exon (Runx1ΔE6) effectively phenocopy the loss of the definitive erythroid lineage in Pcbp2−/−embryos. These data support a model in which enhancement of Runx1 cassette exon 6 inclusion by Pcbp2 serves a critical role in development of hematopoietic progenitors and constitutes a critical step in the developmental pathway of the definitive erythropoietic lineage.

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Pheromone binding protein shapes olfactory temporal resolution

10.1101/2021.01.11.426239 ◽

2021 ◽

Author(s):

Yusuke Shiota ◽

Takeshi Sakurai ◽

Noriyasu Ando ◽

Stephan Shuichi Haupt ◽

Hidefumi Mitsuno ◽

...

Keyword(s):

Temporal Resolution ◽

Binding Protein ◽

Critical Role ◽

High Temporal Resolution ◽

Wild Type ◽

Pheromone Binding Protein ◽

Air Turbulence ◽

Behavioural Analysis ◽

Antennal Response ◽

Pheromone Plumes

AbstractMale moths are capable of orienting towards conspecific females using sex pheromones. Since pheromones are distributed as discontinuous plumes owing to air turbulence, tracking intermittent stimuli with high temporal resolution is suggested to be important for efficient localisation. Here, using a pheromone binding protein (BmPBP1) knockout silkmoth, we revealed that the loss of functional pheromone binding protein altered antennal response kinetics resulting in reduced temporal resolution to intermittent pheromone stimuli on the antennae. Behavioural analysis revealed that BmPBP1-knockout males exhibited significantly less straight walking, which occurs when detecting pheromone stimuli, especially to high frequency stimuli. Accordingly, BmPBP1-knockout males took a significantly longer time to locate pheromone sources and females than did wild-type males. Together, BmPBP1 plays a critical role in determining temporal antennal response kinetics and that an appropriate range of temporal sensory and behavioural resolutions is essential for tracking pheromone plumes for efficient pheromone source localisation in the silkmoth.

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Multivalency enables unidirectional switch-like competition between intrinsically disordered proteins

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2117338119 ◽

2022 ◽

Vol 119 (3) ◽

pp. e2117338119

Author(s):

Rebecca B. Berlow ◽

H. Jane Dyson ◽

Peter E. Wright

Keyword(s):

Transcriptional Control ◽

Intrinsically Disordered Proteins ◽

Binding Protein ◽

Critical Role ◽

Molecular Switch ◽

Negative Regulator ◽

Disordered Proteins ◽

Creb Binding Protein ◽

Binding Motifs ◽

Intrinsically Disordered

Intrinsically disordered proteins must compete for binding to common regulatory targets to carry out their biological functions. Previously, we showed that the activation domains of two disordered proteins, the transcription factor HIF-1α and its negative regulator CITED2, function as a unidirectional, allosteric molecular switch to control transcription of critical adaptive genes under conditions of oxygen deprivation. These proteins achieve transcriptional control by competing for binding to the TAZ1 domain of the transcriptional coactivators CREB-binding protein (CBP) and p300 (CREB: cyclic-AMP response element binding protein). To characterize the mechanistic details behind this molecular switch, we used solution NMR spectroscopy and complementary biophysical methods to determine the contributions of individual binding motifs in CITED2 to the overall competition process. An N-terminal region of the CITED2 activation domain, which forms a helix when bound to TAZ1, plays a critical role in initiating competition with HIF-1α by enabling formation of a ternary complex in a process that is highly dependent on the dynamics and disorder of the competing partners. Two other conserved binding motifs in CITED2, the LPEL motif and an aromatic/hydrophobic motif that we term ϕC, function synergistically to enhance binding of CITED2 and inhibit rebinding of HIF-1α. The apparent unidirectionality of competition between HIF-1α and CITED2 is lost when one or more of these binding regions is altered by truncation or mutation of the CITED2 peptide. Our findings illustrate the complexity of molecular interactions involving disordered proteins containing multivalent interaction motifs and provide insight into the unique mechanisms by which disordered proteins compete for occupancy of common molecular targets within the cell.

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Impaired adipogenesis in insulin-like growth factor binding protein-1 transgenic mice

Journal of Endocrinology ◽

10.1677/joe.0.1620457 ◽

1999 ◽

Vol 162 (3) ◽

pp. 457-465 ◽

Cited By ~ 59

Author(s):

K Rajkumar ◽

T Modric ◽

LJ Murphy

Keyword(s):

Growth Factor ◽

Transgenic Mice ◽

Binding Protein ◽

Critical Role ◽

Insulin Like Growth Factor ◽

Wild Type ◽

Factor Binding ◽

Igf I ◽

Enhanced Expression ◽

Glycerol 3 Phosphate Dehydrogenase

Differentiation of precursor cells into mature fat cells is accompanied by enhanced expression of insulin-like growth factor (IGF)-I and is stimulated by multiple hormones including growth hormone, glucocorticoids, IGF-I and insulin. We used transgenic mice that overexpress insulin-like growth factor binding protein-1 to investigate the role of IGF-I in the accumulation of fat tissue. In response to a sucrose-enriched diet, transgenic mice gained significantly less body weight and the epididymal fat mass was significantly reduced compared with wild-type mice. The increase in adipocyte size was also significantly reduced in transgenic mice compared with wild-type mice. Fewer colonies were generated from adipose tissue from transgenic mice and the mitogenic response of these cells to IGF-I was significantly reduced compared with those from wild-type mice. Induction of glycerol-3-phosphate dehydrogenase, a measure of adipocyte differentiation, by IGF-I but not insulin, was reduced in preadipocytes from transgenic mice. These data indicate that IGF-I has a critical role in the proliferation of adipocyte precursors, the differentiation of preadipocytes and the development of obesity in response to calorie excess.

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MOT1 Can Activate Basal Transcription In Vitro by Regulating the Distribution of TATA Binding Protein between Promoter and Nonpromoter Sites

Molecular and Cellular Biology ◽

10.1128/mcb.19.4.2835 ◽

1999 ◽

Vol 19 (4) ◽

pp. 2835-2845 ◽

Cited By ~ 46

Author(s):

Tamara A. Muldrow ◽

Allyson M. Campbell ◽

P. Anthony Weil ◽

David T. Auble

Keyword(s):

Transcriptional Control ◽

Binding Protein ◽

In Vitro Transcription ◽

Tata Binding Protein ◽

Wild Type ◽

Basal Transcription ◽

Nuclear Extracts ◽

Transcription In Vitro

ABSTRACT MOT1 is an ATPase which can dissociate TATA binding protein (TBP)-DNA complexes in a reaction requiring ATP hydrolysis. Consistent with this observation, MOT1 can repress basal transcription in vitro. Paradoxically, however, some genes, such as HIS4, appear to require MOT1 as an activator of transcription in vivo. To further investigate the function of MOT1 in basal transcription, we performed in vitro transcription reactions using yeast nuclear extracts depleted of MOT1. Quantitation of MOT1 revealed that it is an abundant protein, with nuclear extracts from wild-type cells containing a molar excess of MOT1 over TBP. Surprisingly, MOT1 can weakly activate basal transcription in vitro. This activation by MOT1 is detectable with amounts of MOT1 that are approximately stoichiometric to TBP. With amounts of MOT1 similar to those present in wild-type nuclear extracts, MOT1 behaves as a weak repressor of basal transcription. These results suggest that MOT1 might activate transcription via an indirect mechanism in which limiting TBP can be liberated from nonpromoter sites for use at promoters. In support of this idea, excess nonpromoter DNA sequesters TBP and represses transcription, but this effect can be reversed by addition of MOT1. These results help to reconcile previous in vitro and in vivo results and expand the repertoire of transcriptional control strategies to include factor-assisted redistribution of TBP between promoter and nonpromoter sites.

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MiRNAs and Muscle Regeneration: Therapeutic Targets in Duchenne Muscular Dystrophy

International Journal of Molecular Sciences ◽

10.3390/ijms22084236 ◽

2021 ◽

Vol 22 (8) ◽

pp. 4236

Author(s):

Amelia Eva Aránega ◽

Estefanía Lozano-Velasco ◽

Lara Rodriguez-Outeiriño ◽

Felicitas Ramírez de Acuña ◽

Diego Franco ◽

...

Keyword(s):

Stem Cell ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Muscle Regeneration ◽

Transcriptional Control ◽

Neuromuscular Disorders ◽

Critical Role ◽

Gene Replacement ◽

Control Of Gene Expression ◽

Muscle Disorders

microRNAs (miRNAs) are small non-coding RNAs required for the post-transcriptional control of gene expression. MicroRNAs play a critical role in modulating muscle regeneration and stem cell behavior. Muscle regeneration is affected in muscular dystrophies, and a critical point for the development of effective strategies for treating muscle disorders is optimizing approaches to target muscle stem cells in order to increase the ability to regenerate lost tissue. Within this framework, miRNAs are emerging as implicated in muscle stem cell response in neuromuscular disorders and new methodologies to regulate the expression of key microRNAs are coming up. In this review, we summarize recent advances highlighting the potential of miRNAs to be used in conjunction with gene replacement therapies, in order to improve muscle regeneration in the context of Duchenne Muscular Dystrophy (DMD).

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The DNA Binding Protein Rfg1 Is a Repressor of Filamentation inCandida albicans

Genetics ◽

10.1093/genetics/157.4.1503 ◽

2001 ◽

Vol 157 (4) ◽

pp. 1503-1512 ◽

Cited By ~ 1

Author(s):

Roy A Khalaf ◽

Richard S Zitomer

Keyword(s):

Dna Binding ◽

Binding Protein ◽

Hyphal Growth ◽

Dna Binding Protein ◽

Homozygous Deletion ◽

Wild Type ◽

Pathogenic Yeast ◽

Repression Complex ◽

Type Gene ◽

Repression Domain

AbstractWe have identified a repressor of hyphal growth in the pathogenic yeast Candida albicans. The gene was originally cloned in an attempt to characterize the homologue of the Saccharomyces cerevisiae Rox1, a repressor of hypoxic genes. Rox1 is an HMG-domain, DNA binding protein with a repression domain that recruits the Tup1/Ssn6 general repression complex to achieve repression. The C. albicans clone also encoded an HMG protein that was capable of repression of a hypoxic gene in a S. cerevisiae rox1 deletion strain. Gel retardation experiments using the purified HMG domain of this protein demonstrated that it was capable of binding specifically to a S. cerevisiae hypoxic operator DNA sequence. These data seemed to indicate that this gene encoded a hypoxic repressor. However, surprisingly, when a homozygous deletion was generated in C. albicans, the cells became constitutive for hyphal growth. This phenotype was rescued by the reintroduction of the wild-type gene on a plasmid, proving that the hyphal growth phenotype was due to the deletion and not a secondary mutation. Furthermore, oxygen repression of the hypoxic HEM13 gene was not affected by the deletion nor was this putative ROX1 gene regulated positively by oxygen as is the case for the S. cerevisiae gene. All these data indicate that this gene, now designated RFG1 for Repressor of Filamentous Growth, is a repressor of genes required for hyphal growth and not a hypoxic repressor.

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Recent Progress in the Study of Peroxiredoxin in the Harmful Algal Bloom Species Chattonella marina

Antioxidants ◽

10.3390/antiox10020162 ◽

2021 ◽

Vol 10 (2) ◽

pp. 162

Author(s):

Yohei Shimasaki ◽

Koki Mukai ◽

Yuki Takai ◽

Xuchun Qiu ◽

Yuji Oshima

Keyword(s):

Oxidative Stress ◽

Growth Phase ◽

Critical Role ◽

High Irradiance ◽

Growth Potential ◽

Exponential Growth Phase ◽

Wild Type ◽

Chattonella Marina ◽

Strong Light ◽

Expression Strain

Peroxiredoxin (Prx) is a relatively recently discovered antioxidant enzyme family that scavenges peroxides and is known to be present in organisms from biological taxa ranging from bacteria to multicellular eukaryotes, including photosynthetic organisms. Although there have been many studies of the Prx family in higher plants, green algae, and cyanobacteria, few studies have concerned raphidophytes and dinoflagellates, which are among the eukaryotic algae that cause harmful algal blooms (HABs). In our proteomic study using 2-D electrophoresis, we found a highly expressed 2-Cys peroxiredoxin (2-CysPrx) in the raphidophyte Chattonella marina var. antiqua, a species that induces mass mortality of aquacultured fish. The abundance of the C. marina 2-CysPrx enzyme was highest in the exponential growth phase, during which photosynthetic activity was high, and it then decreased by about a factor of two during the late stationary growth phase. This pattern suggested that 2-CysPrx is a key enzyme involved in the maintenance of high photosynthesis activity. In addition, the fact that the depression of photosynthesis by excessively high irradiance was more severe in the 2-CysPrx low-expression strain (wild type) than in the normal-expression strain (wild type) of C. marina suggested that 2-CysPrx played a critical role in protecting the cell from oxidative stress caused by exposure to excessively high irradiance. In the field of HAB research, estimates of growth potential have been desired to predict the population dynamics of HABs for mitigating damage to fisheries. Therefore, omics approaches have recently begun to be applied to elucidate the physiology of the growth of HAB species. In this review, we describe the progress we have made using a molecular physiological approach to identify the roles of 2-CysPrx and other antioxidant enzymes in mitigating environmental stress associated with strong light and high temperatures and resultant oxidative stress. We also describe results of a survey of expressed Prx genes and their growth-phase-dependent behavior in C. marina using RNA-seq analysis. Finally, we speculate about the function of these genes and the ecological significance of 2-CysPrx, such as its involvement in circadian rhythms and the toxicity of C. marina to fish.

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Stress-induced expression of IPT gene in transgenic wheat reduces grain yield penalty under drought

Journal of Genetic Engineering and Biotechnology ◽

10.1186/s43141-021-00171-w ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Ailin Beznec ◽

Paula Faccio ◽

Daniel J. Miralles ◽

Leonor G. Abeledo ◽

Cecilia Decima Oneto ◽

...

Keyword(s):

Grain Yield ◽

Bread Wheat ◽

Transcriptional Control ◽

Field Conditions ◽

Wild Type ◽

Ipt Gene ◽

Receptor Like Kinase ◽

Biomass Components ◽

Yield Penalty ◽

Wild Type Control

Abstract Background The heterologous expression of isopentenyl transferase (IPT) under the transcriptional control of the senescence-associated receptor-like kinase (SARK) promoter delayed cellular senescence and, through it, increased drought tolerance in plants. To evaluate the effect of pSARK::IPT expression in bread wheat, six independent transgenic events were obtained through the biolistic method and evaluated transgene expression, phenology, grain yield and physiological biomass components in plants grown under both drought and well-irrigating conditions. Experiments were performed at different levels: (i) pots and (ii) microplots inside a biosafety greenhouse, as well as under (iii) field conditions. Results Two transgenic events, called TR1 and TR4, outperformed the wild-type control under drought conditions. Transgenic plants showed higher yield under both greenhouse and field conditions, which was positively correlated to grain number (given by more spikes and grains per spike) than wild type. Interestingly, this yield advantage of the transgenic events was observed under both drought and well-watered conditions. Conclusions The results obtained allow us to conclude that the SARK promoter-regulated expression of the IPT gene in bread wheat not only reduced the yield penalty produced by water stress but also led to improved productivity under well-watered conditions.

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The critical role of T cells in glucocorticoid-induced osteoporosis

Cell Death and Disease ◽

10.1038/s41419-020-03249-4 ◽

2020 ◽

Vol 12 (1) ◽

Author(s):

Lin Song ◽

Lijuan Cao ◽

Rui Liu ◽

Hui Ma ◽

Yanan Li ◽

...

Keyword(s):

Bone Marrow ◽

T Cells ◽

Steady State ◽

Side Effects ◽

Critical Role ◽

Wild Type ◽

Receptor Activator ◽

Steady State Levels ◽

Induced Apoptosis

AbstractGlucocorticoids (GC) are widely used clinically, despite the presence of significant side effects, including glucocorticoid-induced osteoporosis (GIOP). While GC are believed to act directly on osteoblasts and osteoclasts to promote osteoporosis, the detailed underlying molecular mechanism of GC-induced osteoporosis is still not fully elucidated. Here, we show that lymphocytes play a pivotal role in regulating GC-induced osteoporosis. We show that GIOP could not be induced in SCID mice that lack T cells, but it could be re-established by adoptive transfer of splenic T cells from wild-type mice. As expected, T cells in the periphery are greatly reduced by GC; instead, they accumulate in the bone marrow where they are protected from GC-induced apoptosis. These bone marrow T cells in GC-treated mice express high steady-state levels of NF-κB receptor activator ligand (RANKL), which promotes the formation and maturation of osteoclasts and induces osteoporosis. Taken together, these findings reveal a critical role for T cells in GIOP.

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Molecular and Functional Characterization of One Odorant-Binding Protein Gene OBP3 in Bemisia tabaci (Hemiptera: Aleyrodidae)

Journal of Economic Entomology ◽

10.1093/jee/toz248 ◽

2019 ◽

Cited By ~ 1

Author(s):

Ran Wang ◽

Yuan Hu ◽

Peiling Wei ◽

Cheng Qu ◽

Chen Luo

Keyword(s):

Host Plant ◽

Bemisia Tabaci ◽

Binding Protein ◽

Insect Pest ◽

Critical Role ◽

Functional Characterization ◽

Odorant Binding Protein ◽

Binding Characteristics ◽

And Function ◽

Odorant Binding

Abstract Odorant binding proteins (OBPs) of insects play a critical role in chemical perceptions and choice of insect host plant. Bemisia tabaci is a notorious insect pest which can damage more than 600 plant species. In order to explore functions of OBPs in B. tabaci, here we investigated binding characteristics and function of odorant-binding protein 3 in B. tabaci (BtabOBP3). The results indicated that BtabOBP3 shows highly similar sequence with OBPs of other insects, including the typical signature motif of six cysteines. The recombinant BtabOBP3 protein was obtained, and the evaluation of binding affinities to tested volatiles of host plant was conducted, then the results indicated that β-ionone had significantly higher binding to BtabOBP3 among other tested plant volatiles. Furthermore, silencing of BtabOBP3 significantly altered choice behavior of B. tabaci to β-ionone. In conclusion, it has been demonstrated that BtabOBP3 exerts function as one carrier of β-ionone and the results could be contributed to reveal the mechanisms of choosing host plant in B. tabaci.

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