bluuebunny/crossref_metadata_2025_split huggingface dataset:
Clathrin-mediated endocytosis-independent function of VAN3 ARF GTPase Activating Protein at the plasma membrane
Authors: Adamowski Maciek , Friml Jiří 
Published: November 2021

AbstractARF small GTPases in plants serve important cellular functions in subcellular trafficking and developmental functions in auxin-mediated patterning of the plant body. TheArabidopsis thalianaARF regulator ARF-GAP VAN3 has been implicated to act at the plasma membrane (PM) and linked functionally to the clathrin- and dynamin-mediated endocytosis. Here we re-evaluated the localization of VAN3 at the PM and its function in endocytosis. Using Total Internal Reflection Fluorescence microscopy we observed remarkably transient associations of VAN3 to the PM at discrete foci, however, devoid of clathrin, the dynamin isoform DRP1A, or the ARF regulator GNOM, which is also involved in a developmental patterning function mediated from the PM. Clathrin-coated pits are abundant and endocytosis appears to proceed normally invan3-1knockout mutant. In turn, post-translational silencing of clathrin expression indicates that the localization of VAN3 at the PM depends on clathrin function, presumably on clathrin-mediated endocytosis.
The role of clathrin in exocytosis and the mutual regulation of endo- and exocytosis in plant cells
Authors: Adamowski Maciek , Matijević Ivana , Friml Jiří 
Published: November 2021

AbstractWithin the plant endomembrane system, the vesicle coat protein clathrin localizes to the plasma membrane (PM) and thetrans-Golgi Network/Early Endosome (TGN/EE). While the role of clathrin as a major component of endocytosis at the PM is well established, its function at TGN/EE, possibly in exocytosis or the vacuolar pathway, is a matter of debate. This shared function of clathrin also opens a question whether plant cells possess a homeostatic mechanisms that balance rates of opposite trafficking routes, such as endo- and exocytosis. Here we address these questions using lines inducibly silencingCLATHRIN HEAVY CHAIN(CHC). We find a relocation of exocytic soluble and integral membrane protein cargoes to the vacuole, supporting a function of clathrin in exocytosis. A comparison with lines overexpressing AUXILIN-LIKE1, where inhibition of CME precedes rerouting of secretory cargoes, does not support a homeostatic regulatory mechanism adjusting exocytosis to the rates of endocytosis. Complementary experiments reveal only minor and variably detectable reductions in the rates of CME in secretory mutants, also not indicative of a converse homeostatic mechanism adjusting rates of endocytosis to the rates of secretion.
Among retroviral integrases prototype foamy virus integrase displays unique biochemical activities
Authors: Rabe Anthony J. , Tan Yow Yong , Larue Ross C. , Yoder Kristine E. 
Published: November 2021

AbstractIntegrase enzymes of different retroviruses assemble as functional complexes with varying multimers of the protein. Retroviral integrases require a divalent metal cation to perform one-step transesterification catalysis. Tetrameric prototype foamy virus (PFV) intasomes assembled from purified integrase and viral DNA oligonucleotides were characterized for their activity in the presence of different cations. While most retroviral integrases are inactive in calcium, PFV intasomes appear to be uniquely capable of catalysis in calcium. The PFV intasomes also contrast other retroviral integrases by displaying an inverse correlation of activity with increasing manganese beginning at relatively low concentrations. The intasomes were found to be significantly more active in the presence of chloride co-ions compared to acetate. While HIV-1 integrase appears to commit to a target DNA within 20 seconds, PFV intasomes do not commit to target DNA during their reaction lifetime. Together these data highlight the unique biochemical activities of PFV integrase compared to other retroviral integrases.
In vitro culture of bovine fibroblasts using select serum-free media supplemented withChlorella vulgarisextract
Authors: Defendi-Cho Galileo , Gould Timothy M. 
Published: November 2021

AbstractStandard cell culture practices require addition of animal-derived serum to culture media to achieve adequate cell growth. Typically, 5-10% by volume of fetal bovine serum (FBS) is used, which accounts for a vast majority of the cost of media while also imposing environmental and ethical concerns associated with the use of animal serum. Here we tested the efficacy of culturing cells by replacing serum in the media with algae extract and select additives. Using LC-MS, we compared molecular signatures of FBS toChlorellaalgae extracts and identified NAD(H)/NADP(H) as common and relatively abundant features in their characteristic profiles. Bovine fibroblasts, cultured in serum-free media supplemented withC. vulgarisextract and just two growth factors plus insulin, showed significant growth with enhanced viability compared to control cells cultured without serum, albeit still lower than that of controls cultured with 10% FBS. Moreover,C. vulgarisextract enhanced cell viability beyond that of cells cultured with the two growth factors and insulin alone. These results suggest that key components in serum which are essential for cell growth may also be present inC. vulgarisextract, demonstrating that it may be used at least as a partial alternative to serum for cell culture applications.
A Large Database of Chicken Bursal ESTs as a Resource for the Analysis of Vertebrate Gene Function
Authors: Abdrakhmanov Igor , Lodygin Dmitry , Geroth Paul , Arakawa Hiroshi , Law Andy , Plachy Jiri , Korn Berndt , Buerstedde Jean-Marie 
Published: November 2021

Chicken B cells create their immunoglobulin repertoire within the Bursa of Fabricius by gene conversion. The high homologous recombination activity is shared by the bursal B-cell-derived DT40 cell line, which integrates transfected DNA constructs at high rates into its endogenous loci. Targeted integration in DT40 is used frequently to analyze the function of genes by gene disruption. In this paper, we describe a large database of >7000 expressed sequence tags (ESTs) from bursal lymphocytes that should be a valuable resource for the identification of gene disruption targets in DT40. ESTs of interest can be recognized easily by online BLAST or keyword searches. Because the database reflects the gene expression profile of bursal lymphocytes, it provides valuable hints as to which genes might be involved in B-cell–specific processes related to immunoglobulin repertoire formation, signal transduction, transcription, and apoptosis. This large collection of chicken ESTs will also be useful for gene expression studies and comparative gene mapping within the chicken genome project. Details of the bursal EST sequencing project and access to database search forms can be found on the DT40 web site (http://genetics.hpi.uni-hamburg.de/dt40.html).[The sequence data described in this paper have been submitted to the GenBank data library under accession nos. AJ392050–AJ399459.]
MHC-Linked Olfactory Receptor Loci Exhibit Polymorphism and Contribute to Extended HLA/OR-Haplotypes
Authors: Ehlers Anke , Beck Stephan , Forbes Simon A. , Trowsdale John , Volz Armin , Younger Ruth , Ziegler Andreas 
Published: November 2021

Clusters of olfactory receptor (OR) genes are found on most human chromosomes. They are one of the largest mammalian multigene families. Here, we report a systematic study of polymorphism of OR genes belonging to the largest fully sequenced OR cluster. The cluster contains 36 OR genes, of which two belong to the vomeronasal 1 (V1-OR) family. The cluster is divided into a major and a minor region at the telomeric end of the HLA complex on chromosome 6. These OR genes could be involved in MHC-related mate preferences. The polymorphism screen was carried out with 13 genes from the HLA-linked OR cluster and three genes from chromosomes 7, 17, and 19 as controls. Ten human cell lines, representing 18 different chromosome 6s, were analyzed. They were from various ethnic origins and exhibited different HLA haplotypes. All OR genes tested, including those not linked to the HLA complex, were polymorphic. These polymorphisms were dispersed along the coding region and resulted in up to seven alleles for a given OR gene. Three polymorphisms resulted either in stop codons (genes hs6M1-4P,hs6M1-17) or in a 16–bp deletion (gene hs6M1-19P), possibly leading to lack of ligand recognition by the respective receptors in the cell line donors. In total, 13 HLA-linked OR haplotypes could be defined. Therefore, allelic variation appears to be a general feature of human OR genes.[The sequence data reported in this paper have been submitted to EMBL under accession nos. AC006137, AC004178, AJ132194, AL022727, AL031983,AL035402, AL035542, Z98744, CAB55431, AL050339, AL035402, AL096770,AL133267, AL121944, Z98745, AL021808, and AL021807.]
The Mouse Clock Locus: Sequence and Comparative Analysis of 204 Kb from Mouse Chromosome 5
Authors: Wilsbacher Lisa D. , Sangoram Ashvin M. , Antoch Marina P. , Takahashi Joseph S. 
Published: November 2021

The Clock gene encodes a basic helix-loop-helix (bHLH)–PAS transcription factor that regulates circadian rhythms in mice. We previously cloned Clock in mouse and human using a battery of behavioral and molecular techniques, including shotgun sequencing of two bacterial artificial chromosome (BAC) clones. Here we report the finished sequence of a 204-kb region from mouse chromosome 5. This region contains the complete loci for the Clock andTpardl (pFT27) genes, as well as the 3′ partial locus of the Neuromedin U gene; sequence analysis also suggests the presence of two previously unidentified genes. In addition, we provide a comparative genomic sequence analysis with the syntenic region from human chromosome 4. Finally, a new BAC transgenic line indicates that the genomic region that is sufficient for rescue of the Clock mutant phenotype is no greater than 120 kb and tightly flanks the 3′ end of the Clockgene.[The sequence data reported in this paper have been submitted to the GenBank data library under accession no. AF146793.]
Retroposed New Genes Out of the X inDrosophila
Authors: Betrán Esther , Thornton Kevin , Long Manyuan 
Published: November 2021

New genes that originated by various molecular mechanisms are an essential component in understanding the evolution of genetic systems. We investigated the pattern of origin of the genes created by retroposition inDrosophila. We surveyed the wholeDrosophila melanogastergenome for such new retrogenes and experimentally analyzed their functionality and evolutionary process. These retrogenes, functional as revealed by the analysis of expression, substitution, and population genetics, show a surprisingly asymmetric pattern in their origin. There is a significant excess of retrogenes that originate from the X chromosome and retropose to autosomes; new genes retroposed from autosomes are scarce. Further, we found that most of these X-derived autosomal retrogenes had evolved a testis expression pattern. These observations may be explained by natural selection favoring those new retrogenes that moved to autosomes and avoided the spermatogenesis X inactivation, and suggest the important role of genome position for the origin of new genes.[The sequence data from this study have been submitted to GenBank under accession nos.AY150701–AY150797. The following individuals kindly provided reagents, samples, or unpublished information as indicated in the paper: M.-L. Wu, F. Lemeunier, and P. Gibert.]
Comparative Sequence Analysis of Human Minisatellites Showing Meiotic Repeat Instability
Authors: Murray John , Buard Jérôme , Neil David L. , Yeramian Edouard , Tamaki Keiji , Hollies Caroline , Jeffreys Alec J. 
Published: November 2021

The highly variable human minisatellites MS32 (D1S8), MS31A (D7S21), and CEB1 (D2S90) all show recombination-based repeat instability restricted to the germline. Mutation usually results in polar interallelic conversion or occasionally in crossovers, which, at MS32 at least, extend into DNA flanking the repeat array, defining a localized recombination hotspot and suggesting that cis-acting elements in flanking DNA can influence repeat instability. Therefore, comparative sequence analysis was performed to search for common flanking elements associated with these unstable loci. All three minisatellites are located in GC-rich DNA abundant in dispersed and tandem repetitive elements. There were no significant sequence similarities between different loci upstream of the unstable end of the repeat array. Only one of the three loci showed clear evidence for putative coding sequences near the minisatellite. No consistent patterns of thermal stability or DNA secondary structure were shared by DNA flanking these loci. This work extends previous data on the genomic environment of minisatellites. In addition, this work suggests that recombinational activity is not controlled by primary or secondary characteristics of the DNA sequence flanking the repeat array and is not obviously associated with gene promoters as seen in yeast.[The sequence data described in this paper have been submitted to the GenBank data library under accession nos. AF048727(CEB1), AF048728 (MS31A), and AF048729 (MS32).]
Mining SNPs From EST Databases
Authors: Picoult-Newberg Leslie , Ideker Trey E. , Pohl Mark G. , Taylor Scott L. , Donaldson Miriam A. , Nickerson Deborah A. , Boyce-Jacino Michael 
Published: November 2021

There is considerable interest in the discovery and characterization of single nucleotide polymorphisms (SNPs) to enable the analysis of the potential relationships between human genotype and phenotype. Here we present a strategy that permits the rapid discovery of SNPs from publicly available expressed sequence tag (EST) databases. From a set of ESTs derived from 19 different cDNA libraries, we assembled 300,000 distinct sequences and identified 850 mismatches from contiguous EST data sets (candidate SNP sites), without de novo sequencing. Through a polymerase-mediated, single-base, primer extension technique, Genetic Bit Analysis (GBA), we confirmed the presence of a subset of these candidate SNP sites and have estimated the allele frequencies in three human populations with different ethnic origins. Altogether, our approach provides a basis for rapid and efficient regional and genome-wide SNP discovery using data assembled from sequences from different libraries of cDNAs.[The SNPs identified in this study can be found in the National Center of Biotechnology (NCBI) SNP database under submitter handles ORCHID (SNPS-981210-A) and debnick (SNPS-981209-A and SNPS-981209-B).]
A Cluster of ABA-Regulated Genes onArabidopsis thalianaBAC T07M07
Authors: Wang Ming Li , Belmonte Stephen , Kim Ulandt , Dolan Maureen , Morris John W. , Goodman Howard M. 
Published: November 2021

Arabidopsis thalianaBAC T07M07 encoding the abscisic acid-insensitive 4 (ABI4) locus has been sequenced completely. It contains a 95,713-bp insert and 24 predicted genes. Most putative genes were confirmed by gel-based RNA profiling and a cluster of ABA-regulated genes was identified. One of the 24 genes, designatedPP2C5, encodes a putative protein phosphatase 2C. The encoded protein was expressed inEscherichia coli, and its enzyme activity in vitro was confirmed.[The sequence data described in this paper have been submitted to GenBank under accession no.AF085279.]
Efficient Approach to Unique Single-Nucleotide Polymorphism Discovery
Authors: Taillon-Miller Patricia , Piernot Ellen E. , Kwok Pui-Yan 
Published: November 2021

Single-nucleotide polymorphisms (SNPs) are the most frequently found DNA sequence variations in the human genome. It has been argued that a dense set of SNP markers can be used to identify genetic factors associated with complex disease traits. Because all high-throughput genotyping methods require precise sequence knowledge of the SNPs, any SNP discovery approach must involve both the determination of DNA sequence and allele frequencies. Furthermore, high-throughput genotyping also requires a genomic DNA amplification step, making it necessary to develop sequence-tagged sites (STSs) that amplify only the DNA fragment containing the SNP and nothing else from the rest of the genome. In this report, we demonstrate the utility of a SNP-screening approach that yields the DNA sequence and allele frequency information while screening out duplications with minimal cost and effort. Our approach is based on the use of a homozygous complete hydatidiform mole (CHM) as the reference. With this homozygous reference, one can identify and estimate the allele frequencies of common SNPs with a pooled DNA-sequencing approach (rather than having to sequence numerous individuals as is commonly done). More importantly, the CHM reference is preferable to a single individual reference because it reveals readily any duplicated regions of the genome amplified by the PCR assay before the duplicated sequences are found in GenBank. This approach reduces the cost of SNP discovery by 60% and eliminates the costly development of SNP markers that cannot be amplified uniquely from the genome.[Sequence data for this article were deposited with the NCBI dbSTS and dbSNP data libraries under accession nos. G42862–G42905]
The Evolution of Trichromatic Color Vision by Opsin Gene Duplication in New World and Old World Primates
Authors: Dulai Kanwaljit S. , von Dornum Miranda , Mollon John D. , Hunt David M. 
Published: November 2021

Trichromacy in all Old World primates is dependent on separate X-linked MW and LW opsin genes that are organized into a head-to-tail tandem array flanked on the upstream side by a locus control region (LCR). The 5′ regions of these two genes show homology for only the first 236 bp, although within this region, the differences are conserved in humans, chimpanzees, and two species of cercopithecoid monkeys. In contrast, most New World primates have only a single polymorphic X-linked opsin gene; all males are dichromats and trichromacy is achieved only in those females that possess a different form of this gene on each X chromosome. By sequencing the upstream region of this gene in a New World monkey, the marmoset, we have been able to demonstrate the presence of an LCR in an equivalent position to that in Old World primates. Moreover, the marmoset sequence shows extensive homology from the coding region to the LCR with the upstream sequence of the human LW gene, a distance of >3 kb, whereas homology with the human MW gene is again limited to the first 236 bp, indicating that the divergent MW sequence identifies the site of insertion of the duplicated gene. This is further supported by the presence of an incomplete Alu element on the upstream side of this insertion point in the MW gene of both humans and a cercopithecoid monkey, with additional Alu elements present further upstream. Therefore, these Aluelements may have been involved in the initial gene duplication and may also be responsible for the high frequency of gene loss and gene duplication within the opsin gene array. Full trichromacy is present in one species of New World monkey, the howler monkey, in which separate MW and LW genes are again present. In contrast to the separate genes in humans, however, the upstream sequences of the two howler genes show homology with the marmoset for at least 600 bp, which is well beyond the point of divergence of the human MW and LW genes, and each sequence is associated with a different LCR, indicating that the duplication in the howler monkey involved the entire upstream region.[The sequence data described in this paper have been submitted to GenBank under accession nos. AF155218, AF156715, and AF156716.]
Finding New Human Minisatellite Sequences in the Vicinity of Long CA-Rich Sequences
Authors: Giraudeau Fabienne , Petit Elisabeth , Avet-Loiseau Hervé , Hauck Yolande , Vergnaud Gilles , Amarger Valérie 
Published: November 2021

Microsatellites and minisatellites are two classes of tandem repeat sequences differing in their size, mutation processes, and chromosomal distribution. The boundary between the two classes is not defined. We have developed a convenient, hybridization-based human library screening procedure able to detect long CA-rich sequences. Analysis of cosmid clones derived from a chromosome 1 library show that cross-hybridizing sequences tested are imperfect CA-rich sequences, some of them showing a minisatellite organization. All but one of the 13 positive chromosome 1 clones studied are localized in chromosomal bands to which minisatellites have previously been assigned, such as the 1pter cluster. To test the applicability of the procedure to minisatellite detection on a larger scale, we then used a large-insert whole-genome PAC library. Altogether, 22 new minisatellites have been identified in positive PAC and cosmid clones and 20 of them are telomeric. Among the 42 positive PAC clones localized within the human genome by FISH and/or linkage analysis, 25 (60%) are assigned to a terminal band of the karyotype, 4 (9%) are juxtacentromeric, and 13 (31%) are interstitial. The localization of at least two of the interstitial PAC clones corresponds to previously characterized minisatellite-containing regions and/or ancestrally telomeric bands, in agreement with this minisatellite-like distribution. The data obtained are in close agreement with the parallel investigation of human genome sequence data and suggest that long human (CA)s are imperfect CA repeats belonging to the minisatellite class of sequences. This approach provides a new tool to efficiently target genomic clones originating from subtelomeric domains, from which minisatellite sequences can readily be obtained.[The sequence data described in this paper have been submitted to the EMBL data library under accession nos.AJ000377–AJ000383.]
Local thermal non-equilibrium conjugate forced convection and entropy generation in an aircraft cabin with air channel partially filled porous insulation
Authors: Çiçek Oktay , Baytaş A. Cihat 
Published: November 2021

PurposeThe purpose of this study is to numerically investigate heat transfer and entropy generation between airframe and cabin-cargo departments in an aircraft. The conjugate forced convection and entropy generation in a cylindrical cavity within air channel partly filled with porous insulation material as simplified geometry for airframe and cabin-cargo departments are considered under local thermal non-equilibrium condition.Design/methodology/approachThe non-dimensional governing equations for fluid and porous media discretized by finite volume method are solved using the SIMPLE algorithm with pressure and velocity correction.FindingsThe effects of the following parameters on the problem are investigated; Reynolds number, Darcy number, the size of inlet and exit cross-section, thermal conductivity ratio for solid and fluid phases, angle between the vertical symmetry axis and the end of channel wall exit and the gap between adiabatic channel wall and horizontal adiabatic wall separating cabin and cargo sections.Originality/valueThis paper can provide a basic perspective and framework for thermal design between the fuselage and cabin-cargo sections. The minimum total entropy generation number is calculated for various Reynolds numbers and thermal conductivity ratios. It is observed that the channel wall temperature increases for high Reynolds number, low Darcy number, narrower exit cross-section and wider the gap between channel wall and horizontal.