bluuebunny/crossref_metadata_2025 huggingface dataset:
Phase transition between disordered and ordered foraging in Pharaoh's ants
Authors: Beekman Madeleine , Sumpter David J. T. , Ratnieks Francis L. W.
Published: July 2002

The complex collective behavior seen in many insect societies strongly suggests that a minimum number of workers are required for these societies to function effectively. Here we investigated the transition between disordered and ordered foraging in the Pharaoh's ant. We show that small colonies forage in a disorganized manner, with a transition to organized pheromone-based foraging in larger colonies. We also show that when food sources are difficult to locate through independent searching, this transition is first-order and exhibits hysteresis, comparable to a first-order phase transition found in many physical systems. To our knowledge, this is the first experimental evidence of a behavioral phase transition between a maladaptive (disorganized) and an adaptive (organized) state.
BRCA1 mediates ligand-independent transcriptional repression of the estrogen receptor
Authors: Zheng Lei , Annab Lois A. , Afshari Cynthia A. , Lee Wen-Hwa , Boyer Thomas G.
Published: July 2002

Mutational inactivation of BRCA1 confers a cumulative lifetime risk
of breast and ovarian cancers. However, the underlying basis for the
tissue-restricted tumor-suppressive properties of BRCA1 remains poorly
defined. Here we show that BRCA1 mediates ligand-independent
transcriptional repression of the estrogen receptor α (ERα), a
principal determinant of the growth, differentiation, and normal
functional status of breasts and ovaries. In Brca1-null mouse embryo
fibroblasts and BRCA1-deficient human ovarian cancer cells, ERα
exhibited ligand-independent transcriptional activity that was not
observed in Brca1-proficient cells. Ectopic expression in
Brca1-deficient cells of wild-type BRCA1, but not clinically validated
BRCA1 missense mutants, restored ligand-independent repression of ERα
in a manner dependent upon apparent histone deacetylase activity. In
estrogen-dependent human breast cancer cells, chromatin
immunoprecipitation analysis revealed the association of BRCA1 with
ERα at endogenous estrogen-response elements before, but
not after estrogen stimulation. Collectively, these results reveal
BRCA1 to be a ligand-reversible barrier to transcriptional activation
by unliganded promoter-bound ERα and suggest a possible mechanism by
which functional inactivation of BRCA1 could promote
tumorigenesis through inappropriate hormonal regulation of mammary and
ovarian epithelial cell proliferation.
l
-Serine and glycine serve as major astroglia-derived trophic factors for cerebellar Purkinje neurons
Authors: Furuya Shigeki , Tabata Toshihide , Mitoma Junya , Yamada Keiko , Yamasaki Miwako , Makino Asami , Yamamoto Toshifumi , Watanabe Masahiko , Kano Masanobu , Hirabayashi Yoshio
Published: July 2002

Glial cells support the survival and development of central neurons
through the supply of trophic factors. Here we demonstrate that
l
-serine (
l
-Ser) and glycine (Gly) also are
glia-derived trophic factors. These amino acids are released by
astroglial cells and promote the survival, dendritogenesis, and
electrophysiological development of cultured cerebellar Purkinje
neurons. Although
l
-Ser and Gly are generally classified as
nonessential amino acids, 3-phosphoglycerate dehydrogenase (3PGDH), a
key enzyme for their biosynthesis, is not expressed in Purkinje
neurons. By contrast, the Bergman glia, a native astroglia in the
cerebellar cortex, highly expresses 3PGDH. These data suggest that
l
-Ser and Gly mediate the trophic actions of glial cells on
Purkinje neurons.
Polynucleotide phosphorylase functions both as a 3′ → 5′ exonuclease and a poly(A) polymerase in
Escherichia coli
Authors: Mohanty Bijoy K. , Kushner Sidney R.
Published: July 2002

In vitro
, polynucleotide phosphorylase of
Escherichia coli
can both synthesize RNA by using
nucleotide diphosphates as precursors and exonucleolytically degrade
RNA in the presence of inorganic phosphate. However, because of the
high
in vivo
concentration of inorganic phosphate in
exponentially growing cells, it has been assumed that the enzyme works
exclusively as an exonuclease. Here we demonstrate that, contrary to
this prediction, polynucleotide phosphorylase not only synthesizes
long, highly heteropolymeric tails
in vivo
, but also
accounts for all of the observed residual polyadenylylation in poly(A)
polymerase I deficient strains. In addition, the enzyme is responsible
for adding the C and U residues that are found in poly(A) tails in
exponentially growing cultures of wild type
E. coli
.
Cytoplasmic catalytic subunit of protein kinase A mediates cross-repression by NF-κB and the glucocorticoid receptor
Authors: Doucas Vassilis , Shi Yanhong , Miyamoto Shigeki , West Andrew , Verma Inder , Evans Ronald M.
Published: July 2002

Negative transcriptional regulation or cross-coupling between
NF-κB (RelA) and the glucocorticoid receptor (GR) is proposed to play
a regulatory role in human physiology and disease. Despite previous
advances, the biochemical basis of this phenomenon remains a subject of
controversy. We show here that the inhibition of GR activity by RelA
does not require the RelA DNA binding, transactivation, or nuclear
localization domains. Surprisingly, RelA repression of GR is abolished
by mutation of the conserved protein kinase A (PKA) site at amino acid
residue 276 of RelA. We show that GR associates
in vivo
and
in vitro
with the catalytic subunit of PKA (PKAc) in
a ligand-independent manner and that GR transcription depends on PKA
signaling. Indeed, we demonstrated that GR-mediated inhibition of
NF-κB transactivation is PKAc-dependent. In contrast to previous
models, we suggest that the cross-coupling requires a cytoplasmic step
and is regulated by a PKAc-associated signaling.
The human thyrotropin receptor: a heptahelical receptor capable of stimulating members of all four G protein families.
Authors: Laugwitz K L , Allgeier A , Offermanns S , Spicher K , Van Sande J , Dumont J E , Schultz G
Published: July 2002

Thyrotropin is the primary hormone that, via one heptahelical receptor, regulates thyroid cell functions such as secretion, specific gene expression, and growth. In human thyroid, thyrotropin receptor activation leads to stimulation of the adenylyl cyclase and phospholipase C cascades. However, the G proteins involved in thyrotropin receptor action have been only partially defined. In membranes of human thyroid gland, we immunologically identified alpha subunits of the G proteins Gs short, Gs long, Gi1, Gi2, Gi3, G(o) (Go2 and another form of Go, presumably Go1), Gq, G11, G12, and G13. Activation of the thyrotropin (TSH) receptor by bovine TSH led to increased incorporation of the photoreactive GTP analogue [alpha-32P]GTP azidoanilide into immunoprecipitated alpha subunits of all G proteins detected in thyroid membranes. This effect was receptor-dependent and not due to direct G protein stimulation because it was mimicked by TSH receptor-stimulating antibodies of patients suffering from Grave disease and was abolished by a receptor-blocking antiserum from a patient with autoimmune hypothyroidism. The TSH-induced activation of individual G proteins occurred with EC50 values of 5-50 milliunits/ml, indicating that the activated TSH receptor coupled with similar potency to different G proteins. When human thyroid slices were pretreated with pertussis toxin, the TSH receptor-mediated accumulation of cAMP increased by approximately 35% with TSH at 1 milliunits/ml, indicating that the TSH receptor coupled to Gs and G(i). Taken together, these findings show that, at least in human thyroid membranes, in which the protein is expressed at its physiological levels, the TSH receptor resembles a naturally occurring example of a general G protein-activating receptor.
The whistle and the rattle: the design of sound producing muscles.
Authors: Rome L C , Syme D A , Hollingworth S , Lindstedt S L , Baylor S M
Published: July 2002

Vertebrate sound producing muscles often operate at frequencies exceeding 100 Hz, making them the fastest vertebrate muscles. Like other vertebrate muscle, these sonic muscles are "synchronous," necessitating that calcium be released and resequestered by the sarcoplasmic reticulum during each contraction cycle. Thus to operate at such high frequencies, vertebrate sonic muscles require extreme adaptations. We have found that to generate the "boatwhistle" mating call (approximately 200 Hz), the swimbladder muscle fibers of toadfish have evolved (i) a large and very fast calcium transient, (ii) a fast crossbridge detachment rate, and (iii) probably a fast kinetic off-rate of Ca2+ from troponin. The fibers of the shaker muscle of rattlesnakes have independently evolved similar traits, permitting tail rattling at approximately 90 Hz.
Down-regulation of transcripts for Na channel α-SNS in spinal sensory neurons following axotomy
Authors: Dib-Hajj S. , Black J. A. , Felts P. , Waxman S. G.
Published: July 2002

Spinal sensory (dorsal root ganglion; DRG) neurons display slowly
inactivating, tetrodotoxin-resistant (TTX-R), and rapidly inactivating,
TTX-sensitive (TTX-S) Na currents. Attenuation of the TTX-R Na current
and enhancement of TTX-S Na current have been demonstrated in cutaneous
afferent DRG neurons in the adult rat after axotomy and may underlie
abnormal bursting. We show here that steady-state levels of transcripts
encoding the α-SNS subunit, which is associated with a slowly
inactivating, TTX-R current when expressed in oocytes, are reduced
significantly 5 days following axotomy of DRG neurons, and continue to
be expressed at reduced levels, even after 210 days. Steady-state
levels of α-III transcripts, which are present at low levels in
control DRG neurons, show a pattern of transiently increased
expression.
In situ
hybridization using α-SNS- and
α-III-specific riboprobes showed a decreased signal for α-SNS, and
an increased signal for α-III, in both large and small DRG neurons
following axotomy. Reduced levels of α-SNS may explain the selective
loss of slowly inactivating, TTX-R current. The abnormal
electrophysiological properties of DRG neurons following axonal injury
thus appear to reflect a switch in Na channel gene expression.
Superior efficacy of secreted over somatic antigen display in recombinant Salmonella vaccine induced protection against listeriosis.
Authors: Hess J , Gentschev I , Miko D , Welzel M , Ladel C , Goebel W , Kaufmann S H
Published: July 2002

Vaccination provides the most potent measure against infectious disease, and recombinant (r) viable vaccines expressing defined pathogen-derived antigens represent powerful candidates for future vaccination strategies. In a new approach we constructed r-aroA- Salmonella typhimurium displaying p60 or listeriolysin (Hly) antigen of Listeria monocytogenes in secreted or somatic form in the host cell. Vaccination of mice with r-aroA- S. typhimurium induced protection against the intracellular pathogen L. monocytogenes only with secreted and not with somatic antigen. Secreted Hly was slightly more potent in inducing protective immunity than secreted p60. Both r-aroA- S. typhimurium secreting p60 in the endosome and r-aroA- S. typhimurium secreting Hly in the cytosol induced protective CD4+ and CD8+ T-cells suggesting CD8+ T-cell stimulation independent from intracellular residence of r-aroA- S. typhimurium carriers. Hence, not only the type of antigen but also its display by the r-carrier within the host cell critically influences vaccine efficacy.
Antisense globin RNA in mouse erythroid tissues: structure, origin, and possible function.
Authors: Volloch V , Schweitzer B , Rits S
Published: July 2002

The aim of the experiments described in this paper was to test for the presence of antisense globin RNA in mouse erythroid tissues and, if found, to characterize these molecules. The present study made use of a multistep procedure in which a molecular tag is attached to cellular RNA by ligation with a defined ribooligonucleotide. The act of ligation preserves the termini of RNA molecules, which become the junctions between cellular RNAs and the ligated ribooligonucleotide. It also unambiguously preserves the identity of cellular RNA as a sense or antisense molecule through all subsequent manipulations. Using this approach, we identified and characterized antisense beta-globin RNA in erythroid spleen cells and reticulocytes from anemic mice. We show in this paper that the antisense globin RNA is fully complementary to spliced globin mRNA, indicative of the template/transcript relationship. It terminates at the 5' end with a uridylate stretch, reflecting the presence of poly(A) at the 3' end of the sense globin mRNA. With respect to the structure of their 3' termini, antisense globin RNA can be divided into three categories: full-size molecules corresponding precisely to globin mRNA, truncated molecules lacking predominantly 14 3'-terminal nucleotides, and extended antisense RNA containing 17 additional 3'-terminal nucleotides. The full-size antisense globin RNA contains two 14-nt-long complementary sequences within its 3'-terminal segment corresponding to the 5'-untranslated region of globin mRNA. This, together with the nature of the predominant truncation, suggests a mechanism by which antisense RNA might give rise to new sense-strand globin mRNA.
Implications of fault constitutive properties for earthquake prediction.
Authors: Dieterich J H , Kilgore B
Published: July 2002

The rate- and state-dependent constitutive formulation for fault slip characterizes an exceptional variety of materials over a wide range of sliding conditions. This formulation provides a unified representation of diverse sliding phenomena including slip weakening over a characteristic sliding distance Dc, apparent fracture energy at a rupture front, time-dependent healing after rapid slip, and various other transient and slip rate effects. Laboratory observations and theoretical models both indicate that earthquake nucleation is accompanied by long intervals of accelerating slip. Strains from the nucleation process on buried faults generally could not be detected if laboratory values of Dc apply to faults in nature. However, scaling of Dc is presently an open question and the possibility exists that measurable premonitory creep may precede some earthquakes. Earthquake activity is modeled as a sequence of earthquake nucleation events. In this model, earthquake clustering arises from sensitivity of nucleation times to the stress changes induced by prior earthquakes. The model gives the characteristic Omori aftershock decay law and assigns physical interpretation to aftershock parameters. The seismicity formulation predicts large changes of earthquake probabilities result from stress changes. Two mechanisms for foreshocks are proposed that describe observed frequency of occurrence of foreshock-mainshock pairs by time and magnitude. With the first mechanism, foreshocks represent a manifestation of earthquake clustering in which the stress change at the time of the foreshock increases the probability of earthquakes at all magnitudes including the eventual mainshock. With the second model, accelerating fault slip on the mainshock nucleation zone triggers foreshocks.
Cooperative functions of the
reaper
and
head involution defective
genes in the programmed cell death of
Drosophila
central nervous system midline cells
Authors: Zhou L. , Schnitzler A. , Agapite J. , Schwartz L. M. , Steller H. , Nambu J. R.
Published: July 2002

In
Drosophila
, the chromosomal region 75C1–2 contains at least three genes,
reaper
(
rpr
),
head involution defective
(
hid
), and
grim
, that have important functions in the activation of programmed cell death. To better understand how cells are killed by these genes, we have utilized a well defined set of embryonic central nervous system midline cells that normally exhibit a specific pattern of glial cell death. In this study we show that both
rpr
and
hid
are expressed in dying midline cells and that the normal pattern of midline cell death requires the function of multiple genes in the 75C1–2 interval. We also utilized the P[UAS]/P[Gal4] system to target expression of
rpr
and
hid
to midline cells. Targeted expression of
rpr
or
hid
alone was not sufficient to induce ectopic midline cell death. However, expression of both
rpr
and
hid
together rapidly induced ectopic midline cell death that resulted in axon scaffold defects characteristic of mutants with abnormal midline cell development. Midline-targeted expression of the baculovirus p35 protein, a caspase inhibitor, blocked both normal and ectopic
rpr
- and
hid
-induced cell death. Taken together, our results suggest that
rpr
and
hid
are expressed together and cooperate to induce programmed cell death during development of the central nervous system midline.
G
sα
contains an unidentified covalent modification that increases its affinity for adenylyl cyclase
Authors: Kleuss Christiane , Gilman Alfred G.
Published: July 2002

Many G protein α subunits are dually acylated with myristate and palmitate or are palmitoylated on more than one cysteine residue near their N termini. The G
α
protein that activates adenylyl cyclase, α
s
, is not myristoylated but can be reversibly palmitoylated. It appears that α
s
contains another, as-yet-unidentified covalent modification that decreases its apparent dissociation constant for adenylyl cyclase from 50 nM to <0.5 nM. This modification is at or near the N terminus of the protein and is hydrophobic. Palmitoylation of native α
s
does not account for its high affinity for adenylyl cyclase.
Evolution of HIV-1 coreceptor usage through interactions with distinct CCR5 and CXCR4 domains
Authors: Lu Zhao-hai , Berson Joanne F. , Chen Ying-hua , Turner Julie D. , Zhang Tian-yuan , Sharron Matthew , Jenks M. Harley , Wang Zi-xuan , Kim Jin , Rucker Joseph , Hoxie James A. , Peiper Stephen C. , Doms Robert W.
Published: July 2002

The chemokine receptor CXCR4 functions as a fusion coreceptor for T cell tropic and dual-tropic HIV-1 strains. To identify regions of CXCR4 that are important for coreceptor function, CXCR4–CXCR2 receptor chimeras were tested for the ability to support HIV-1 envelope (env) protein-mediated membrane fusion. Receptor chimeras containing the first and second extracellular loops of CXCR4 supported fusion by T tropic and dual-tropic HIV-1 and HIV-2 strains and binding of a monoclonal antibody to CXCR4, 12G5, that blocks CXCR4-dependent infection by some virus strains. The second extracellular loop of CXCR4 was sufficient to confer coreceptor function to CXCR2 for most virus strains tested but did not support binding of 12G5. Truncation of the CXCR4 cytoplasmic tail or mutation of a conserved DRY motif in the second intracellular loop did not affect coreceptor function, indicating that phosphorylation of the cytoplasmic tail and the DRY motif are not required for coreceptor function. The results implicate the involvement of multiple CXCR4 domains in HIV-1 coreceptor function, especially the second extracellular loop, though the structural requirements for coreceptor function were somewhat variable for different env proteins. Finally, a hybrid receptor in which the amino terminus of CXCR4 was replaced by that of CCR5 was active as a coreceptor for M tropic, T tropic, and dual-tropic env proteins. We propose that dual tropism may evolve in CCR5-restricted HIV-1 strains through acquisition of the ability to utilize the first and second extracellular loops of CXCR4 while retaining the ability to interact with the CCR5 amino-terminal domain.
MLL is fused to CBP, a histone acetyltransferase, in therapy-related acute myeloid leukemia with a t(11;16)(q23;p13.3)
Authors: Sobulo Olatoyosi M. , Borrow Julian , Tomek Ronald , Reshmi Shalini , Harden Alanna , Schlegelberger Brigitte , Housman David , Doggett Norman A. , Rowley Janet D. , Zeleznik-Le Nancy J.
Published: July 2002

The recurring translocation t(11;16)(q23;p13.3) has been documented only in cases of acute leukemia or myelodysplasia secondary to therapy with drugs targeting DNA topoisomerase II. We show that the
MLL
gene is fused to the gene that codes for CBP (CREB-binding protein), the protein that binds specifically to the DNA-binding protein CREB (cAMP response element-binding protein) in this translocation.
MLL
is fused in-frame to a different exon of
CBP
in two patients producing chimeric proteins containing the AT-hooks, methyltransferase homology domain, and transcriptional repression domain of MLL fused to the CREB binding domain or to the bromodomain of CBP. Both fusion products retain the histone acetyltransferase domain of CBP and may lead to leukemia by promoting histone acetylation of genomic regions targeted by the MLL AT-hooks, leading to transcriptional deregulation via aberrant chromatin organization.
CBP
is the first partner gene of
MLL
containing well defined structural and functional motifs that provide unique insights into the potential mechanisms by which these translocations contribute to leukemogenesis.
Functional characterization of the C—C chemokine-like molecules encoded by molluscum contagiosum virus types 1 and 2
Authors: Krathwohl Mitchell D. , Hromas Robert , Brown Darron R. , Broxmeyer Hal E. , Fife Kenneth H.
Published: July 2002

Many viruses have evolved mechanisms for evading the host immune system by synthesizing proteins that interfere with the normal immune response. The poxviruses are among the most accomplished at deceiving their hosts’ immune systems. The nucleotide sequence of the genome of the human cutaneous poxvirus, molluscum contagiosum virus (MCV) type 1, was recently reported to contain a region that resembles a human chemokine. We have cloned and expressed the chemokine-like genes from MCV type 1 and the closely related MCV type 2 to determine a potential role for these proteins in the viral life cycle. In monocyte chemotaxis assays, the viral proteins have no chemotactic activity but both viral proteins block the chemotactic response to the human chemokine, macrophage inflammatory protein (MIP)-1α. Like MIP-1α, both viral proteins also inhibit the growth of human hematopoietic progenitor cells, but the viral proteins are more potent in this activity than the human chemokine. These viral chemokines antagonize the chemotactic activity of human chemokines and have an inhibitory effect on human hematopoietic progenitor cells. We hypothesize that the inhibition of chemotaxis is an immune evasion function of these proteins during molluscum contagiosum virus infection. The significance of hematopoietic progenitor cell inhibition in viral pathogenesis is uncertain.
Leukemia-associated retinoic acid receptor α fusion partners, PML and PLZF, heterodimerize and colocalize to nuclear bodies
Authors: Koken M. H. M. , Reid A. , Quignon F. , Chelbi-Alix M. K. , Davies J. M. , Kabarowski J. H. S. , Zhu J. , Dong S. , Chen S.-J. , Chen Z. , Tan C. C. , Licht J. , Waxman S. , de Thé H. , Zelent A.
Published: July 2002

In acute promyelocytic leukemia (APL), the typical t(15;17) and the rare t(11;17) translocations express, respectively, the PML/RARα and PLZF/RARα fusion proteins (where RARα is retinoic acid receptor α). Herein, we demonstrate that the PLZF and PML proteins interact with each other and colocalize onto nuclear bodies (NBs). Furthermore, induction of PML expression by interferons leads to a recruitment of PLZF onto NBs without increase in the levels of the PLZF protein. PML/RARα and PLZF/RARα localize to the same microspeckled nuclear domains that appear to be common targets for the two fusion proteins in APL. Although PLZF/RARα does not affect the localization of PML, PML/RARα delocalizes the endogenous PLZF protein in t(15;17)-positive NB4 cells, pointing to a hierarchy in the nuclear targeting of these proteins. Thus, our results unify the molecular pathogenesis of APL with at least two different RARα gene translocations and stress the importance of alterations of PLZF and RARα nuclear localizations in this disease.
Dopamine-induced recruitment of dopamine D1 receptors to the plasma membrane
Authors: Brismar Hjalmar , Asghar Mohammad , Carey Robert M. , Greengard Paul , Aperia Anita
Published: July 2002

The recruitment of G protein-coupled receptors from the cytoplasm to the plasma membrane generally is believed to be a constitutive process. We show here by the use of both confocal microscopy and subcellular fractionation that, for at least one such receptor, this recruitment is regulated and not constitutive. Cells from a proximal tubular-like cell line, LLCPK1 cells, were incubated with either a D1 agonist, a dopamine precursor, or an inhibitor of dopamine metabolism to increase dopamine availability in the cell. Each of the three procedures led to a rapid translocation of dopamine D1 receptors from the cytosol to the plasma membrane.
Regulation of the human p21/WAF1/Cip1 promoter in hepatic cells by functional interactions between Sp1 and Smad family members
Authors: Moustakas Aristidis , Kardassis Dimitris
Published: July 2002

The cell cycle inhibitor p21/WAF1/Cip1 is expressed in many cell types and is regulated by p53-dependent and p53-independent mechanisms. p21 is an important regulator of hepatocyte cell cycle, differentiation, and liver development, but little is known about the regulation of its synthesis in hepatocytes. We report herein that the p21 gene is constitutively expressed in human hepatoma HepG2 cells. Deletion analysis of the p21 promoter showed that it contains a distal (positions −2,300/−210) and a proximal (positions −124 to −61) region that act synergistically to achieve high levels of constitutive expression. The proximal region that consists of multiple Sp1 binding sites is essential for constitutive p21 promoter activity in hepatocytes. This region also mediates the transcriptional activation of the p21 promoter by members of the Smad family of proteins, which play important role in the transduction of extracellular signals such as transforming growth factor β, activin, etc. Constitutive expression of p21 was severely reduced by a C-terminally truncated form of Smad4 that was shown previously to block signaling through Smads. Smad3/4 and to a much lesser extent Smad2/4 caused high levels of transcriptional activation of the p21 promoter. Transactivation was compromised by N- or C-terminally truncated forms of Smad3. By using Gal4-Sp1 fusion proteins, we show that Smad proteins can activate gene transcription via functional interactions with the ubiquitous factor Sp1. These data demonstrate that Smad proteins and Sp1 participate in the constitutive or inducible expression of the p21 gene in hepatic cells.
Productivity, individual-level and colony-level flexibility, and organization of work as consequences of colony size
Authors: Karsai István , Wenzel John W.
Published: July 2002

In social insects, colony-level complexity may emerge from simple individual-level behaviors and interactions. Emergent global properties such as colony size, which can be viewed as a consequence of life history traits, may influence individual-level behaviors themselves. The effects of colony size on productivity, body size, behavioral flexibility, and colony organization are examined here by considering colony size as an independent variable. Large colony size commonly corresponds with complex colony-level performance, small body size, and lower per capita productivity. Analyzing the construction behavior of various wasp societies reveals that complexity of individual behavior is inversely related to colony size. Parallel processing by specialists in large colonies provides flexible and efficient colony-level functioning. On the other hand, individual behavioral flexibility of jack-of-all trades workers ensures success of the small and early societies.
The
BRCA
2
gene product functionally interacts with p53 and RAD51
Authors: Marmorstein Lihua Y. , Ouchi Toru , Aaronson Stuart A.
Published: July 2002

Germ-line mutations in the human
BRCA2
gene confer susceptibility to breast cancer. Efforts to elucidate its function have revealed a putative transcriptional activation domain and
in vitro
interaction with the DNA repair protein RAD51. Other studies have indicated that RAD51 physically associates with the p53 tumor suppressor protein. Here we show that the
BRCA2
gene product is a 460-kDa nuclear phosphoprotein, which forms
in vivo
complexes with both p53 and RAD51. Moreover, exogenous BRCA2 expression in cancer cells inhibits p53’s transcriptional activity, and RAD51 coexpression enhances BRCA2’s inhibitory effects. These findings demonstrate that BRCA2 physically and functionally interacts with two key components of cell cycle control and DNA repair pathways. Thus, BRCA2 likely participates with p53 and RAD51 in maintaining genome integrity.
Formation of the preprimosome protects λ O from RNA transcription-dependent proteolysis by ClpP/ClpX
Authors: Zylicz Maciej , Liberek Krzysztof , Wawrzynow Alicja , Georgopoulos Costa
Published: July 2002

Using the bacteriophage λ DNA replication system, composed entirely of purified proteins, we have tested the accessibility of the short-lived λ O protein to the ClpP/ClpX protease during the various stages of λ DNA replication. We find that binding of λ O protein to its
ori
λ DNA sequence, leading to the so-called “O-some” formation, largely inhibits its degradation. On the contrary, under conditions permissive for transcription, the λ O protein bound to the
ori
λ sequence becomes largely accessible to ClpP/ClpX-mediated proteolysis. However, when the λ O protein is part of the larger
ori
λ:O⋅P⋅DnaB preprimosomal complex, transcription does not significantly increase ClpP/ClpX-dependent λ O degradation. These results show that transcription can stimulate proteolysis of a protein that is required for the initiation of DNA replication.
Hematopoietic stem cell deficiencies in mice lacking c-Mpl, the receptor for thrombopoietin
Authors: Kimura Shinya , Roberts Andrew W. , Metcalf Donald , Alexander Warren S.
Published: July 2002

Thrombopoietin (TPO) acts as a lineage-specific late-acting factor to stimulate megakaryocyte and platelet formation. However, analysis of mice lacking either the cytokine or its receptor, c-Mpl, also revealed deficiencies in progenitor cells of multiple hematopoietic lineages, suggesting that TPO signaling may play an important role in the regulation of the hematopoietic stem cell compartment. To investigate this hypothesis, we determined preprogenitor and colony forming unit-spleen (CFU-S) numbers and analyzed the long-term hematopoietic repopulating capacity of bone marrow cells from
mpl
−/−
mice.
mpl
−/−
mice had 4- to 12-fold fewer preprogenitor cells than wild-type mice. In irradiated normal recipients,
mpl
−/−
bone marrow generated 8- to 10-fold fewer spleen colonies than wild-type marrow at both 8 and 12 days after transplantation. This defect was intrinsic to the transplanted hematopoietic cells, as the microenvironment of
mpl
−/−
recipients supported similar CFU-S growth to that observed in wild-type recipients. In definitive assays of stem cell function, bone marrow cells from
mpl
−/−
mice failed to compete effectively with normal cells for long-term reconstitution of the hematopoietic organs of irradiated recipients, even when transplanted in 10-fold excess. Serial transplantation studies further suggested that stem cell self-renewal also may be compromised in
mpl
−/−
mice. These data imply that TPO, signaling through c-Mpl, plays a vital physiological role in the regulation of hematopoietic stem cell production and function.
Crystal structure of the pleckstrin homology-phosphotyrosine binding (PH-PTB) targeting region of insulin receptor substrate 1
Authors: Dhe-Paganon Sirano , Ottinger Elizabeth A. , Nolte Robert T. , Eck Michael J. , Shoelson Steven E.
Published: July 2002

We have determined the crystal structure at 2.3-Å resolution of an amino-terminal segment of human insulin receptor substrate 1 that encompasses its pleckstrin homology (PH) and phosphotyrosine binding (PTB) domains. Both domains adopt the canonical seven-stranded β-sandwich PH domain fold. The domains are closely associated, with a 720-Å
2
contact surface buried between them that appears to be stabilized by ionic, hydrophobic, and hydrogen bonding interactions. The nonconserved 46-residue linker between the domains is disordered. The PTB domain peptide binding site is fully exposed on the molecular surface, as is a large cationic patch at the base of the PH domain that is a likely binding site for the head groups of phosphatidylinositol phosphates. Binding assays confirm that phosphatidylinositol phosphates bind the PH domain, but not the PTB domain. Ligand binding to the PH domain does not alter PTB domain interactions, and vice versa. The structural and accompanying functional data illustrate how the two binding domains might act cooperatively to effectively increase local insulin receptor substrate 1 concentration at the membrane and transiently fix the receptor and substrate, to allow multiple phosphorylation reactions to occur during each union.
Amplification and overexpression of peroxisome proliferatoractivated receptor binding protein (
PBP/PPARBP
) gene in breast cancer
Authors: Zhu Yijun , Qi Chao , Jain Sanjay , Le Beau Michelle M. , Espinosa Rafael , Atkins G. Brandon , Lazar Mitchell A. , Yeldandi Anjana V. , Rao M. Sambasiva , Reddy Janardan K.
Published: July 2002

Peroxisome proliferator-activated receptor binding protein (PBP), a nuclear receptor coactivator, interacts with estrogen receptor α (ERα) in the absence of estrogen. This interaction was enhanced in the presence of estrogen but was reduced in the presence of antiestrogen, tamoxifen. Transfection of PBP in CV-1 cells resulted in enhancement of estrogen-dependent transcription, indicating that PBP serves as a coactivator in ER signaling. To examine whether overexpression of PBP plays a role in breast cancer because of its coactivator function in ER signaling, we determined the levels of PBP expression in breast tumors. High levels of PBP expression were detected in ≈50% of primary breast cancers and breast cancer cell lines by ribonuclease protection analysis,
in situ
hybridization, and immunoperoxidase staining. Fluorescence
in situ
hybridization of human chromosomes revealed that the
PBP
gene is located on chromosome 17q12, a region that is amplified in some breast cancers. We found
PBP
gene amplification in ≈24% (6/25) of breast tumors and ≈30% (2/6) of breast cancer cell lines, implying that PBP gene overexpression can occur independent of gene amplification. This gene comprises 17 exons that, together, span >37 kilobases. The 5′-flanking region of 2.5 kilobase pairs inserted into a luciferase reporter vector revealed that the promoter activity in CV-1 cells increased by deletion of nucleotides from −2,500 to −273. The −273 to +1 region, which exhibited high promoter activity, contains a typical CCAT box and multiple cis-elements such as C/EBPβ, YY1, c-Ets-1, AP1, AP2, and NFκB binding sites. These observations, in particular
PBP
gene amplification, suggest that PBP, by its ability to function as ERα coactivator, might play a role in mammary epithelial differentiation and in breast carcinogenesis.
Heme is an effector molecule for iron-dependent degradation of the bacterial iron response regulator (Irr) protein
Authors: Qi Zhenhao , Hamza Iqbal , O’Brian Mark R.
Published: July 2002

The bacterial iron response regulator (Irr) protein mediates iron-dependent regulation of heme biosynthesis. Pulse–chase and immunoprecipitation experiments showed that Irr degraded in response to 6 μM iron with a half-life of ≈30 min and that this regulated stability was the principal determinant of control by iron. Irr contains a heme regulatory motif (HRM) near its amino terminus. A role for heme in regulation was implicated by the retention of Irr in heme synthesis mutants in the presence of iron. Addition of heme to low iron (0.3 μM) cultures was sufficient for the disappearance of Irr in cells of the wild-type and heme mutant strains. Spectral and binding analyses of purified recombinant Irr showed that the protein bound heme with high affinity and caused a blue shift in the absorption spectrum of heme to a shorter wavelength. A Cys
29
→ Ala substitution within the HRM of Irr (IrrC29A) abrogated both high affinity binding to heme and the spectral blue shift.
In vivo
turnover experiments showed that, unlike wild-type Irr, IrrC29A was stable in the presence of iron. We conclude that iron-dependent degradation of Irr involves direct binding of heme to the protein at the HRM. The findings implicate a regulatory role for heme in protein degradation and provide direct evidence for a functional HRM in a prokaryote.
Unique forms of human and mouse nuclear receptor corepressor SMRT
Authors: Ordentlich Peter , Downes Michael , Xie Wen , Genin Anna , Spinner Nancy B. , Evans Ronald M.
Published: July 2002

Nuclear hormone receptors have been shown to repress transcription in the absence of ligand. This repression is mediated by a corepressor complex that contains the Sin3A protein and histone deacetylases (HDAC1 and 2). Studies by several groups demonstrate that this complex is recruited to nuclear receptors through the highly related corepressors SMRT (silencing mediator of retinoid acid and thyroid hormone receptor) and N-CoR (nuclear receptor corepressor). We describe here the cloning, characterization, and chromosomal mapping of forms of human and mouse SMRT that includes a 1,000-aa extension, which reveals striking homology to the amino terminus of N-CoR. Structure and function studies of wild-type and natural splicing variants suggest the presence of 3–4 amino terminal domains that repress in a cooperative as well as mechanistically distinct fashion.
Glutathione synthesis is essential for mouse development but not for cell growth in culture
Authors: Shi Zheng-Zheng , Osei-Frimpong Joseph , Kala Geeta , Kala Subbarao V. , Barrios Roberto J. , Habib Geetha M. , Lukin Dana J. , Danney Christopher M. , Matzuk Martin M. , Lieberman Michael W.
Published: July 2002

Glutathione (GSH) is a major source of reducing equivalents in mammalian cells. To examine the role of GSH synthesis in development and cell growth, we generated mice deficient in GSH by a targeted disruption of the heavy subunit of γ-glutamylcysteine synthetase (γGCS-HStm1), an essential enzyme in GSH synthesis. Embryos homozygous for γGCS-HStm1fail to gastrulate, do not form mesoderm, develop distal apoptosis, and die before day 8.5. Lethality results from apoptotic cell death rather than reduced cell proliferation. We also isolated cell lines from homozygous mutant blastocysts in medium containing GSH. These cells also grow indefinitely in GSH-free medium supplemented withN-acetylcysteine and have undetectable levels of GSH; further, they show no changes in mitochondrial morphology as judged by electron microscopy. These data demonstrate that GSH is required for mammalian development but dispensable in cell culture and that the functions of GSH, not GSH itself, are essential for cell growth.
tmRNAs that encode proteolysis-inducing tags are found in all known bacterial genomes: A two-piece tmRNA functions in
Caulobacter
Authors: Keiler Kenneth C. , Shapiro Lucy , Williams Kelly P.
Published: July 2002

A general mechanism in bacteria to rescue stalled ribosomes and to clear the cell of incomplete polypeptides involves an RNA species, tmRNA (SsrA), which functions as both a tRNA and an mRNA. This RNA encodes a peptide tag that is incorporated at the end of the aberrant polypeptide and targets it for proteolysis. We have identified a circularly permuted version of the tmRNA gene in α-proteobacteria as well as in a lineage of cyanobacteria. The genes in these two groups seem to have arisen from two independent permutation events. As a result of the altered genetic structure, these tmRNAs are composed of two distinct RNA molecules. The mature two-piece tmRNAs are predicted to have a tRNA-like domain and an mRNA-like domain similar to those of standard one-piece tmRNAs, with a break located in the loop containing the tag reading frame. A related sequence was found in the mitochondrial genome of
Reclinomonas americana
, but only the tRNA-like portion is retained. Although several sequence and structural motifs that are conserved among one-piece tmRNAs have been lost, the α-proteobacterium
Caulobacter crescentus
produces a functional two-piece tmRNA.
m
5
C RNA and m
5
C DNA methyl transferases use different cysteine residues as catalysts
Authors: Liu Yaoquan , Santi Daniel V.
Published: July 2002

A family of RNA m
5
C methyl transferases (MTases) containing over 55 members in eight subfamilies has been identified recently by an iterative search of the genomic sequence databases by using the known 16S rRNA m
5
C 967 MTase, Fmu, as an initial probe. The RNA m
5
C MTase family contained sequence motifs that were highly homologous to motifs in the DNA m
5
C MTases, including the ProCys sequence that contains the essential Cys catalyst of the functionally similar DNA-modifying enzymes; it was reasonable to assign the Cys nucleophile to be that in the conserved ProCys. The family also contained an additional conserved Cys residue that aligns with the nucleophilic catalyst in m
5
U54 tRNA MTase. Surprisingly, the mutant of the putative Cys catalyst in the ProCys sequence was active and formed a covalent complex with 5-fluorocytosine-containing RNA, whereas the mutant at the other conserved Cys was inactive and unable to form the complex. Thus, notwithstanding the highly homologous sequences and similar functions, the RNA m
5
C MTase uses a different Cys as a catalytic nucleophile than the DNA m
5
C MTases. The catalytic Cys seems to be determined, not by the target base that is modified, but by whether the substrate is DNA or RNA. The function of the conserved ProCys sequence in the RNA m
5
C MTases remains unknown.
Hydrolytic editing by a class II aminoacyl-tRNA synthetase
Authors: Beuning Penny J. , Musier-Forsyth Karin
Published: July 2002

Editing reactions catalyzed by aminoacyl-tRNA synthetases are critical for accurate translation of the genetic code. To date, this activity, whereby misactivated amino acids are hydrolyzed either before or after transfer to noncognate tRNAs, has been characterized extensively only in the case of class I synthetases. Class II synthetases have an active-site architecture that is completely distinct from that of class I. Thus, findings on editing by class I synthetases may not be applicable generally to class II enzymes. Class II
Escherichia coli
proline-tRNA synthetase is shown here to misactivate alanine and to hydrolyze the noncognate amino acid before transfer to tRNA
Pro
. This enzyme also is capable of rapidly deacylating a mischarged Ala-tRNA
Pro
variant. A single cysteine residue (C443) that is located within the class II-specific motif 3 consensus sequence was shown previously to be dispensable for proline-tRNA synthetase aminoacylation activity. We show here that C443 is critical for the hydrolytic editing of Ala-tRNA
Pro
by this class II synthetase.
BAR: An apoptosis regulator at the intersection of caspases and Bcl-2 family proteins
Authors: Zhang Hong , Xu Qunli , Krajewski Stanislaw , Krajewska Maryla , Xie Zhihua , Fuess Sally , Kitada Shinichi , Pawłowski Krzysztof , Godzik Adam , Reed John C.
Published: July 2002

Two major pathways for induction of apoptosis have been identified—intrinsic and extrinsic. The extrinsic pathway is represented by tumor necrosis factor family receptors, which utilize protein interaction modules known as death domains and death effector domains (DEDs) to assemble receptor signaling complexes that recruit and activate certain caspase-family cell death proteases, namely procaspases-8 and -10. The intrinsic pathway for apoptosis involves the participation of mitochondria, which release caspase-activating proteins. Bcl-2 family proteins govern this mitochondria-dependent apoptosis pathway, with proteins such as Bax functioning as inducers and proteins such as Bcl-2 and Bcl-X
L
serving as suppressors of cell death. An apoptosis regulator, BAR, was identified by using a yeast-based screen for inhibitors of Bax-induced cell death. The BAR protein contains a SAM domain, which is required for its interactions with Bcl-2 and Bcl-X
L
and for suppression of Bax-induced cell death in both mammalian cells and yeast. In addition, BAR contains a DED-like domain responsible for its interaction with DED-containing procaspases and suppression of Fas-induced apoptosis. Furthermore, BAR can bridge procaspase-8 and Bcl-2 into a protein complex. The BAR protein is anchored in intracellular membranes where Bcl-2 resides. BAR therefore may represent a scaffold protein capable of bridging two major apoptosis pathways.
Monitoring CD8 T cell responses to NY-ESO-1: Correlation of humoral and cellular immune responses
Authors: Jäger Elke , Nagata Yasuhiro , Gnjatic Sacha , Wada Hisashi , Stockert Elisabeth , Karbach Julia , Dunbar P. Rod , Lee Sang Yull , Jungbluth Achim , Jäger Dirk , Arand Michael , Ritter Gerd , Cerundolo Vincenzo , Dupont Bo , Chen Yao-Tseng , Old Lloyd J. , Knuth Alexander
Published: July 2002

NY-ESO-1, a member of the cancer–testis family of antigens, is expressed in a subset of a broad range of different human tumor types. Patients with advanced NY-ESO-1-expressing tumors frequently develop humoral immunity to NY-ESO-1, and three HLA A2-restricted peptides were defined previously as targets for cytotoxic CD8+T cells in a melanoma patient with NY-ESO-1 antibody. The objectives of the present study were (i) to develop enzyme-linked immunospot (ELISPOT) and tetramer assays to measure CD8+T cell responses to NY-ESO-1, (ii) to determine the frequency of CD8+T cell responses to NY-ESO-1 in a series of HLA-A2 patients with NY-ESO-1 expressing tumors, (iii) to determine the relation between CD8+T cell and humoral immune responses to NY-ESO-1, and (iv) to compare results of NY-ESO-1 ELISPOT assays performed independently in two laboratories with T cells from the same patients. NY-ESO-1 ELISPOT and tetramer assays with excellent sensitivity, specificity, and reproducibility have been developed and found to correlate with cytotoxicity assays. CD8+T cell responses to HLA-A2-restricted NY-ESO-1 peptides were detected in 10 of 11 patients with NY-ESO-1 antibody, but not in patients lacking antibody or in patients with NY-ESO-1-negative tumors. The results of ELISPOT assays were concordant in the two laboratories, providing the basis for standardized monitoring of T cell responses in patients receiving NY-ESO-1 vaccines.