bluuebunny/crossref_metadata_2025_split huggingface dataset:
Introns and Splicing Elements of Five Diverse Fungi
Authors: Kupfer Doris M. , Drabenstot Scott D. , Buchanan Kent L. , Lai Hongshing , Zhu Hua , Dyer David W. , Roe Bruce A. , Murphy Juneann W. 
Published: October 2004

ABSTRACT

Genomic sequences and expressed sequence tag data for a diverse group of fungi (
Saccharomyces cerevisiae
,
Schizosaccharomyces pombe
,
Aspergillus nidulans
,
Neurospora crassa
, and
Cryptococcus neoformans
) provided the opportunity to accurately characterize conserved intronic elements. An examination of large intron data sets revealed that fungal introns in general are short, that 98% or more of them belong to the canonical splice site (ss) class (5′GU…AG3′), and that they have polypyrimidine tracts predominantly in the region between the 5′ ss and the branch point. Information content is high in the 5′ ss, branch site, and 3′ ss regions of the introns but low in the exon regions adjacent to the introns in the fungi examined. The two yeasts have broader intron length ranges and correspondingly higher intron information content than the other fungi. Generally, as intron length increases in the fungi, so does intron information content. Homologs of U2AF spliceosomal proteins were found in all species except for
S. cerevisiae
, suggesting a nonconventional role for U2AF in the absence of canonical polypyrimidine tracts in the majority of introns. Our observations imply that splicing in fungi may be different from that in vertebrates and may require additional proteins that interact with polypyrimidine tracts upstream of the branch point. Theoretical protein homologs for Nam8p and TIA-1, two proteins that require U-rich regions upstream of the branch point to function, were found. There appear to be sufficient differences between
S. cerevisiae
and
S. pombe
introns and the introns of two filamentous members of the Ascomycota and one member of the Basidiomycota to warrant the development of new model organisms for studying the splicing mechanisms of fungi.
Transcriptional Transitions during
Dictyostelium
Spore Germination
Authors: Xu Qikai , Ibarra Miroslava , Mahadeo Dana , Shaw Chad , Huang Eryong , Kuspa Adam , Cotter David , Shaulsky Gad 
Published: October 2004

ABSTRACT

Many protozoa form spores in response to adversity; therefore, spore germination is a key process in their life cycle.
Dictyostelium discoideum
sporulates in response to starvation following a developmental program. Germination is characterized by two visible changes, spore swelling and the emergence of amoeba from the spore capsule. Several studies have indicated that an additional process termed spore activation is also required, but the physiological changes that characterize the three phases are largely uncharacterized. We used microarrays to monitor global transcriptional transitions as a surrogate measure of the physiological changes that occur during germination. Using two independent methods to induce germination, we identified changes in mRNA levels that characterized the germination process rather than changes that resulted from the induction method. We found that germination is characterized by three transitions. The first transition occurs during activation, while the spores appear dormant, the largest transition occurs when swelling begins, and the third transition occurs when emergence begins. These findings indicate that activation and swelling are not passive occurrences, such as dilution of inhibitors or spore rehydration, but are active processes that are accompanied by dramatic events in mRNA degradation and de novo transcription. These findings confirm and extend earlier reports that genes such as
celA
are regulated during spore germination. We also found by mutation analysis that the unconventional myosin gene
myoI
, which is induced during early germination, plays roles in the maintenance of dormancy and in spore swelling. This finding suggests that some of the observed transcriptional changes are required for spore germination.
Msn2- and Msn4-Like Transcription Factors Play No Obvious Roles in the Stress Responses of the Fungal Pathogen
Candida albicans
Authors: Nicholls Susan , Straffon Melissa , Enjalbert Brice , Nantel André , Macaskill Susan , Whiteway Malcolm , Brown Alistair J. P. 
Published: October 2004

ABSTRACT

In
Saccharomyces cerevisiae
, the (C
2
H
2
)
2
zinc finger transcription factors Msn2 and Msn4 play central roles in responses to a range of stresses by activating gene transcription via the stress response element (STRE; CCCCT). The pathogen
Candida albicans
displays stress responses that are thought to help it survive adverse environmental conditions encountered within its human host. However, these responses differ from those in
S. cerevisiae
, and hence we predicted that the roles of Msn2- and Msn4-like proteins might have been functionally reassigned in
C. albicans. C. albicans
has two such proteins: CaMsn4 and Mnl1 (for Msn2- and Msn4-like).
CaMSN4
, but not
MNL1
, weakly complemented the inability of an
S. cerevisiae msn2 msn4
mutant to activate a STRE
-lacZ
reporter. Also, the disruption of CaMsn4 and Mnl1 had no discernible effect upon the resistance of
C. albicans
to heat, osmotic, ethanol, nutrient, oxidative, or heavy-metal stress or upon the stress-activated transcriptome in
C. albicans
. Furthermore, although Cap1-dependent activation of a Yap response element-luciferase reporter was observed, a STRE reporter was not activated in response to stresses in
C. albicans
. Ectopic expression of CaMsn4 or Mnl1 did not affect the cellular or molecular responses of
C. albicans
to stress. Under the conditions tested, the putative activation and DNA binding domains of
CaMsn4
did not appear to be functional. These data suggest that CaMsn4 and Mnl1 do not contribute significantly to stress responses in
C. albicans
. The data are consistent with the idea that stress signaling in this fungus has diverged significantly from that in budding yeast.
Integration of Stress Responses: Modulation of Calcineurin Signaling in
Saccharomyces cerevisiae
by Protein Kinase A
Authors: Kafadar Kimberly A. , Cyert Martha S. 
Published: October 2004

ABSTRACT

Calcineurin is a Ca
2+
/calmodulin-dependent protein phosphatase required for
Saccharomyces cerevisiae
to adapt to a variety of environmental stresses. Once activated, calcineurin dephosphorylates the Zn-finger transcription factor Crz1p/Tcn1p, causing it to accumulate in the nucleus where it activates gene expression. Here we show that cyclic AMP-dependent protein kinase A (PKA) phosphorylates and negatively regulates Crz1p activity by inhibiting its nuclear import. Activation of PKA in vivo decreases Crz1p-dependent transcription. PKA phosphorylates Crz1p in vitro, and we identify specific residues required for this phosphorylation, all of which reside in or adjacent to the nuclear localization signal. Mutation of these residues to alanine results in increased nuclear import of Crz1p and results in higher levels of both basal and Ca
2+
-induced Crz1p transcriptional activity. PKA regulates the general stress response in yeast and coordinates this response with nutrient availability. In contrast, calcineurin regulates the cellular response to a restricted set of environmental insults. Thus, these studies identify a specific biochemical mechanism through which the activities of multiple stress-activated signaling pathways are integrated in vivo.
Inner Kinetochore of the Pathogenic YeastCandida glabrata
Authors: Stoyan Tanja , Carbon John 
Published: October 2004

ABSTRACTThe human pathogenic yeastCandida glabratais the second most commonCandidapathogen afterCandida albicans, causing both bloodstream and mucosal infections. The centromere (CEN) DNA ofC. glabrata(CgCEN), although structurally very similar to that ofSaccharomyces cerevisiae, is not functional inS. cerevisiae. To further examine the structure of theC. glabratainner kinetochore, we isolated severalC. glabratahomologs ofS. cerevisiaeinner kinetochore protein genes, namely, genes for components of the CBF3 complex (Ndc10p, Cep3p, and Ctf13p) and genes for the proteins Mif2p and Cse4p. The amino acid sequence identities of these proteins were 32 to 49% relative toS. cerevisiae. CgNDC10,CgCEP3, andCgCTF13are required for growth inC. glabrataand are specifically found atCgCEN, as demonstrated by chromatin immunoprecipitation experiments. Cross-complementation experiments revealed that the isolated genes, with the exception ofCgCSE4, are species specific and cannot functionally substitute for the corresponding genes inS. cerevisiaedeletion strains. Likewise, theS. cerevisiaeCBF3 genesNDC10,CEP3, andCTF13cannot functionally replace their homologs inC. glabrataCBF3 deletion strains. Two-hybrid analysis revealed several interactions between these proteins, all of which were previously reported for the inner kinetochore proteins ofS. cerevisiae. Our findings indicate that although many of the inner kinetochore components have evolved considerably between the two closely related species, the organization of theC. glabratainner kinetochore is similar to that inS. cerevisiae.
Gene Replacement of Fructose-1,6-Bisphosphate Aldolase Supports the Hypothesis of a Single Photosynthetic Ancestor of Chromalveolates
Authors: Patron Nicola J. , Rogers Matthew B. , Keeling Patrick J. 
Published: October 2004

ABSTRACT
Plastids (photosynthetic organelles of plants and algae) are known to have spread between eukaryotic lineages by secondary endosymbiosis, that is, by the uptake of a eukaryotic alga by another eukaryote. But the number of times this has taken place is controversial. This is particularly so in the case of eukaryotes with plastids derived from red algae, which are numerous and diverse. Despite their diversity, it has been suggested that all these eukaryotes share a recent common ancestor and that their plastids originated in a single endosymbiosis, the so-called “chromalveolate hypothesis.” Here we describe a novel molecular character that supports the chromalveolate hypothesis. Fructose-1,6-bisphosphate aldolase (FBA) is a glycolytic and Calvin cycle enzyme that exists as two nonhomologous types, class I and class II. Red algal plastid-targeted FBA is a class I enzyme related to homologues from plants and green algae, and it would be predicted that the plastid-targeted FBA from algae with red algal secondary endosymbionts should be related to this class I enzyme. However, we show that plastid-targeted FBA of heterokonts, cryptomonads, haptophytes, and dinoflagellates (all photosynthetic chromalveolates) are class II plastid-targeted enzymes, completely unlike those of red algal plastids. The chromalveolate enzymes form a strongly supported group in FBA phylogeny, and their common possession of this unexpected plastid characteristic provides new evidence for their close relationship and a common origin for their plastids.
A Cell Number Counting Factor Regulates Akt/Protein Kinase B To Regulate
Dictyostelium discoideum
Group Size
Authors: Gao Tong , Knecht David , Tang Lei , Hatton R. Diane , Gomer Richard H. 
Published: October 2004

ABSTRACT

Little is known about how individual cells can organize themselves to form structures of a given size. During development,
Dictyostelium discoideum
aggregates in dendritic streams and forms groups of ∼20,000 cells.
D. discoideum
regulates group size by secreting and simultaneously sensing a multiprotein complex called counting factor (CF). If there are too many cells in a stream, the associated high concentration of CF will decrease cell-cell adhesion and increase cell motility, causing aggregation streams to break up. The pulses of cyclic AMP (cAMP) that mediate aggregation cause a transient translocation of Akt/protein kinase B (Akt/PKB) to the leading edge of the plasma membrane and a concomitant activation of the kinase activity, which in turn stimulates motility. We found that

countin
−

cells (which lack bioactive CF) and wild-type cells starved in the presence of anticountin antibodies (which block CF activity) showed a decreased level of cAMP-stimulated Akt/PKB membrane translocation and kinase activity compared to parental wild-type cells. Recombinant countin has the bioactivity of CF, and a 1-min treatment of cells with recombinant countin potentiated Akt/PKB translocation to membranes and Akt/PKB activity. Western blotting of total cell lysates indicated that countin does not affect the total level of Akt/PKB. Fluorescence microscopy of cells expressing an Akt/PKB pleckstrin homology domain-green fluorescent protein (PH-GFP) fusion protein indicated that recombinant countin and anti-countin antibodies do not obviously alter the distribution of Akt/PKB PH-GFP when it translocates to the membrane. Our data indicate that CF increases motility by potentiating the cAMP-stimulated activation and translocation of Akt/PKB.
Nucleus-Encoded Genes for Plastid-Targeted Proteins in
Helicosporidium
: Functional Diversity of a Cryptic Plastid in a Parasitic Alga
Authors: de Koning Audrey P. , Keeling Patrick J. 
Published: October 2004

ABSTRACT

Plastids are the organelles of plants and algae that house photosynthesis and many other biochemical pathways. Plastids contain a small genome, but most of their proteins are encoded in the nucleus and posttranslationally targeted to the organelle. When plants and algae lose photosynthesis, they virtually always retain a highly reduced “cryptic” plastid. Cryptic plastids are known to exist in many organisms, although their metabolic functions are seldom understood. The best-studied example of a cryptic plastid is from the intracellular malaria parasite,
Plasmodium
, which has retained a plastid for the biosynthesis of fatty acids, isoprenoids, and heme by the use of plastid-targeted enzymes. To study a completely independent transformation of a photosynthetic plastid to a cryptic plastid in another alga-turned-parasite, we conducted an expressed sequence tag (EST) survey of
Helicosporidium
. This parasite has recently been recognized as a highly derived green alga. Based on phylogenetic relationships to other plastid homologues and the presence of N-terminal transit peptides, we have identified 20 putatively plastid-targeted enzymes that are involved in a wide variety of metabolic pathways. Overall, the metabolic diversity of the
Helicosporidium
cryptic plastid exceeds that of the
Plasmodium
plastid, as it includes representatives of most of the pathways known to operate in the
Plasmodium
plastid as well as many others. In particular, several amino acid biosynthetic pathways have been retained, including the leucine biosynthesis pathway, which was only recently recognized in plant plastids. These two parasites represent different evolutionary trajectories in plastid metabolic adaptation.
TbDSS-1, an Essential
Trypanosoma brucei
Exoribonuclease Homolog That Has Pleiotropic Effects on Mitochondrial RNA Metabolism
Authors: Penschow Jonelle L. , Sleve Daniel A. , Ryan Christopher M. , Read Laurie K. 
Published: October 2004

ABSTRACT

Mitochondrial gene expression in trypanosomes is controlled primarily at the levels of RNA processing and RNA stability. This regulation undoubtedly involves numerous ribonucleases. Here we characterize the
Trypanosoma brucei
homolog of the yeast DSS-1 mitochondrial exoribonuclease, which we term TbDSS-1. Biochemical fractionation indicates that TbDSS-1 is mitochondrially localized, as predicted by its N-terminal sequence. In contrast to its yeast homolog, TbDSS-1 does not appear to be associated with mitochondrial ribosomes. Targeted downregulation of TbDSS-1 by RNA interference in procyclic-form
T. brucei
results in a severe growth defect. In addition, TbDSS-1 depletion leads to a decrease in the levels of never edited cytochrome oxidase subunit I (COI) mRNA and both unedited and edited COIII mRNAs, indicating this enzyme functions in the control of mitochondrial RNA abundance. We also observe a considerable reduction in the level of edited apocytochrome b (CYb) mRNA and a corresponding increase in unedited CYb mRNA, suggesting that TbDSS-1 functions, either directly or indirectly, in the control of RNA editing. The abundance of both gCYb[560] and gA6[149] guide RNAs is reduced upon TbDSS-1 depletion, although the reduction in gCYb[560] is much more dramatic. The significant reduction in gCYb levels could potentially account for the observed decrease in CYb RNA editing. Western blot analyses of mitochondrial RNA editing and stability factors indicate that the perturbations of RNA levels observed in TbDSS-1 knock-downs do not result from secondary effects on other mitochondrial proteins. In all, these data demonstrate that TbDSS-1 is an essential protein that plays a role in mitochondrial RNA stability and RNA editing.
Functional Characterization of Myosin I Tail Regions in
Candida albicans
Authors: Oberholzer Ursula , Iouk Tatiana L. , Thomas David Y. , Whiteway Malcolm 
Published: October 2004

ABSTRACT

The molecular motor myosin I is required for hyphal growth in the pathogenic yeast
Candida albicans
. Specific myosin I functions were investigated by a deletion analysis of five neck and tail regions. Hyphal formation requires both the TH1 region and the IQ motifs. The TH2 region is important for optimal hyphal growth. All of the regions, except for the SH3 and acidic (A) regions that were examined individually, were required for the localization of myosin I at the hyphal tip. Similarly, all of the domains were required for the association of myosin I with pelletable actin-bound complexes. Moreover, the hyphal tip localization of cortical actin patches, identified by both rhodamine-phalloidin staining and Arp3-green fluorescent protein signals, was dependent on myosin I. Double deletion of the A and SH3 domains depolarized the distribution of the cortical actin patches without affecting the ability of the mutant to form hyphae, suggesting that myosin I has distinct functions in these processes. Among the six myosin I tail domain mutants, the ability to form hyphae was strictly correlated with endocytosis. We propose that the uptake of cell wall remodeling enzymes and excess plasma membrane is critical for hyphal formation.
FKBP12 Controls Aspartate Pathway Flux in
Saccharomyces cerevisiae
To Prevent Toxic Intermediate Accumulation
Authors: Arévalo-Rodríguez Miguel , Pan Xuewen , Boeke Jef D. , Heitman Joseph 
Published: October 2004

ABSTRACT

FKBP12 is a conserved member of the prolyl-isomerase enzyme family and serves as the intracellular receptor for FK506 that mediates immunosuppression in mammals and antimicrobial actions in fungi. To investigate the cellular functions of FKBP12 in
Saccharomyces cerevisiae
, we employed a high-throughput assay to identify mutations that are synthetically lethal with a mutation in the
FPR1
gene, which encodes FKBP12. This screen identified a mutation in the
HOM6
gene, which encodes homoserine dehydrogenase, the enzyme catalyzing the last step in conversion of aspartic acid into homoserine, the common precursor in threonine and methionine synthesis. Lethality of
fpr1 hom6
double mutants was suppressed by null mutations in
HOM3
or
HOM2
, encoding aspartokinase and aspartate β-semialdehyde dehydrogenase, respectively, supporting the hypothesis that
fpr1 hom6
double mutants are inviable because of toxic accumulation of aspartate β-semialdehyde, the substrate of homoserine dehydrogenase. Our findings also indicate that mutation or inhibition of FKBP12 dysregulates the homoserine synthetic pathway by perturbing aspartokinase feedback inhibition by threonine. Because this pathway is conserved in fungi but not in mammals, our findings suggest a facile route to synergistic antifungal drug development via concomitant inhibition of FKBP12 and Hom6.
Insights into the Survival of
Chlamydomonas reinhardtii
during Sulfur Starvation Based on Microarray Analysis of Gene Expression
Authors: Zhang Zhaoduo , Shrager Jeff , Jain Monica , Chang Chiung-Wen , Vallon Olivier , Grossman Arthur R. 
Published: October 2004

ABSTRACT

Responses of photosynthetic organisms to sulfur starvation include (i) increasing the capacity of the cell for transporting and/or assimilating exogenous sulfate, (ii) restructuring cellular features to conserve sulfur resources, and (iii) modulating metabolic processes and rates of cell growth and division. We used microarray analyses to obtain a genome-level view of changes in mRNA abundances in the green alga
Chlamydomonas reinhardtii
during sulfur starvation. The work confirms and extends upon previous findings showing that sulfur deprivation elicits changes in levels of transcripts for proteins that help scavenge sulfate and economize on the use of sulfur resources. Changes in levels of transcripts encoding members of the light-harvesting polypeptide family, such as LhcSR2, suggest restructuring of the photosynthetic apparatus during sulfur deprivation. There are also significant changes in levels of transcripts encoding enzymes involved in metabolic processes (e.g., carbon metabolism), intracellular proteolysis, and the amelioration of oxidative damage; a marked and sustained increase in mRNAs for a putative vanadium chloroperoxidase and a peroxiredoxin may help prolong survival of
C. reinhardtii
during sulfur deprivation. Furthermore, many of the sulfur stress-regulated transcripts (encoding polypeptides associated with sulfate uptake and assimilation, oxidative stress, and photosynthetic function) are not properly regulated in the
sac1
mutant of
C. reinhardtii
, a strain that dies much more rapidly than parental cells during sulfur deprivation. Interestingly, sulfur stress elicits dramatic changes in levels of transcripts encoding putative chloroplast-localized chaperones in the
sac1
mutant but not in the parental strain. These results suggest various strategies used by photosynthetic organisms during acclimation to nutrient-limited growth.
All-terrain vehicle injury in children: strategies for prevention: Table 1
Authors: Aitken M E , Graham C J , Killingsworth J B , Mullins S H , Parnell D N , Dick R M 
Published: October 2004

Objective:A variety of educational efforts, policies, and regulations have been adopted to reduce all-terrain vehicle (ATV) injury in children. Despite this, ATV use by children continues and serious injuries are common. The purpose of this study was to investigate the knowledge, practices, and beliefs of ATV users to help develop effective educational strategies to promote safer ATV use.Design:Focus groups were conducted to characterize participant ATV use and safety awareness as well as to explore avenues for prevention. Feedback on draft ATV safety public service announcements was elicited. Themes of transcribed focus group data were summarized.Setting:Rural state with high ATV use and injury rates.Subjects:Adult and adolescent ATV users.Interventions:None.Main outcome measures:Summaries of focus group discussions.Results:ATV riders frankly discussed current use and safety behaviors and were aware of some ATV risks. Youths felt that age specific regulation was unlikely to be a helpful strategy. Participants endorsed messages demonstrating graphic consequences as likely to get the attention of young riders regarding risks. Educational settings were suggested, including hunter and driver safety classes.Conclusions:Efforts to improve ATV safety awareness should clearly show pediatric ATV injury risk and safety practices. Campaigns must also show realistic understanding of current use practices to be credible for users. Messages emphasizing the consequences of ATV use were endorsed as most likely to have impact. Approaches based on age based restrictions were considered unrealistic and alternative strategies were suggested.