bluuebunny/crossref_metadata_2025 huggingface dataset:
Genes involved in Haemophilus influenzae type b capsule expression are part of an 18-kilobase tandem duplication.
Authors: Hoiseth S K , Moxon E R , Silver R P
Published: May 2006

Encapsulated Haemophilus influenzae type b produce nonencapsulated variants at high frequency (0.1-0.3%). Cosmid cloning was used to investigate the genetic mechanism responsible for this instability. Analysis of three independently derived cosmid clones showed that the b+ parental strain contains an 18-kilobase tandem duplication of genes involved in type b capsule expression. Loss of one complete copy of the 18-kilobase tandem duplication occurred following transformation of the cosmid clones into Rec+, but not Rec-, Escherichia coli, and in H. influenzae strains that had spontaneously lost capsule expression. These results suggest that high-frequency loss of type b capsule expression is due to rec-dependent recombination between the two copies of the 18-kilobase tandem repeat. This is further supported by our finding that introduction of the H. influenzae rec-1 mutation stabilized type b capsule expression.
Extinction of Harrington's mountain goat
Authors: Mead Jim I. , Martin Paul S. , Euler Robert C. , Long Austin , Jull A. J. T. , Toolin Laurence J. , Donahue Douglas J. , Linick T. W.
Published: May 2006

Keratinous horn sheaths of the extinct Harrington's mountain goat,
Oreamnos harringtoni
, were recovered at or near the surface of dry caves of the Grand Canyon, Arizona. Twenty-three separate specimens from two caves were dated nondestructively by the tandem accelerator mass spectrometer (TAMS). Both the TAMS and the conventional dates indicate that Harrington's mountain goat occupied the Grand Canyon for at least 19,000 years prior to becoming extinct by 11,160 ± 125 radiocarbon years before present. The youngest average radiocarbon dates on Shasta ground sloths,
Nothrotheriops shastensis
, from the region are not significantly younger than those on extinct mountain goats. Rather than sequential extinction with Harrington's mountain goat disappearing from the Grand Canyon before the ground sloths, as one might predict in view of evidence of climatic warming at the time, the losses were concurrent. Both extinctions coincide with the regional arrival of Clovis hunters.
On the relationship among three theories of relaxation in disordered systems
Authors: Klafter Joseph , Shlesinger Michael F.
Published: May 2006

Empirical evidence has accumulated showing the wide occurrence of a stretched-exponential relaxation decay law for many diverse condensed-matter systems. Several theories based on different physical mechanisms have been successful in deriving the stretched-exponential decay law in a natural way. Three of these theories, the direct-transfer model, the hierarchically constrained dynamics model, and the defect-diffusion model are shown here to have an underlying common mathematical structure.
Inhibition of splicing but not cleavage at the 5' splice site by truncating human beta-globin pre-mRNA.
Authors: Furdon P J , Kole R
Published: May 2006

Human beta-globin mRNAs truncated in the second exon or in the first intron have been processed in vitro in a HeLa cell nuclear extract. Transcripts containing a fragment of the second exon as short as 53 nucleotides are efficiently spliced, whereas transcripts truncated 24 or 14 nucleotides downstream from the 3' splice site are spliced inefficiently, if at all. All of these transcripts, however, are efficiently and accurately cleaved at the 5' splice site. In contrast, RNA truncated in the first intron, 54 nucleotides upstream from the 3' splice site, is not processed at all. These findings suggest that cleavage at the 5' splice site and subsequent splicing steps--i.e., cleavage at the 3' splice site and exon ligation--need not be coupled. Anti-Sm serum inhibits the complete splicing reaction and cleavage at the 5' splice site, suggesting involvement of certain ribonucleoprotein particles in the cleavage reaction. ATP and Mg2+ are required for cleavage at the 5' splice site at concentrations similar to those for the complete splicing reaction.
Cytoplasmic synthesis of globin RNA in differentiated murine erythroleukemia cells: possible involvement of RNA-dependent RNA polymerase.
Authors: Volloch V
Published: May 2006

Three lines of evidence indicate that RNA-dependent RNA synthesis occurs in mouse erythroleukemia cells. The first involves labeling studies with [3H]uridine and shows a greater initial labeling rate of globin RNA in the cytoplasm than in the nucleus. Labeled globin RNA found in the cytoplasm after a very short pulse with tritiated uridine is of the "mature" 9S size while labeled globin RNA in the nuclei is exclusively in the form of 15S precursor molecules, suggesting that cytoplasmic globin RNA is not of nuclear origin. A high concentration of actinomycin D has no effect on the initial rate of labeling of cytoplasmic globin RNA, supporting this conclusion. Other experiments showed that the labeling of cytoplasmic globin RNA does not involve end addition to preexisting globin RNA. The second line of evidence is the identification of globin RNA minus strand in the cytoplasm of differentiated murine erythroleukemia cells by hybridization with single-stranded DNA probes containing the strand of the same sense as globin mRNA. This material has the same electrophoretic mobility as globin RNA and hybridizes with probes containing only the 5' part or only the 3' part of the gene suggesting that it is a full size copy of globin RNA. Finally, in murine erythroleukemia cells an RNA-dependent RNA polymerase activity is detected by using poly(A) . oligo(U) as a template-primer combination. This activity increases significantly after induction, suggesting that it is differentiation specific.
Earthworms produce a collagen-like substance detected by the garter snake vomeronasal system.
Authors: Kirschenbaum D M , Schulman N , Halpern M
Published: May 2006

Earthworms (Lumbricus terrestris) produce a chemical substance that is readily detected by and serves as an attractant for garter snakes (Thamnophis sirtalis). This chemoattractant is sensed by the vomeronasal system of snakes. Amino acid analysis of the chemoattractant revealed a high hydroxyproline/proline ratio and large amounts of serine and threonine. More than one-third of the residues were glycine. No hydroxylysine and no cysteine were present. Carbohydrate analyses revealed a high content of galactose (11% by weight) and smaller amounts of fucose, mannose, glucose, N-acetylglucosamine, and N-acetylgalactosamine. These results were very similar to results reported for the amino acid composition and carbohydrate content of earthworm cuticle collagen and gelatin. A preparation of purified earthworm cuticle collagen proved to be a potent source of chemoattractant for garter snakes. Further, it was not possible to prepare chemoattractant from decuticlized earthworms. These results strongly suggest that a component of the earthworm chemoattractant for snakes is structurally related to earthworm cuticle collagen.
The primary structure of rat parathymosin.
Authors: Komiyama T , Pan L X , Haritos A A , Wideman J W , Pan Y C , Chang M , Rogers I , Horecker B L
Published: May 2006

Parathymosin has been isolated from rat thymus and from rat liver. Its primary structure is reported as follows: (Sequence; see text). The blocking group at the NH2 terminus was identified by mass spectrometry as acetyl. Regions homologous to amino acid sequences in prothymosin alpha were found to be located between residues 14-20, 23-25, 33-39, 41-43, and 83-87 of parathymosin.
Metastable photoproducts from carbon monoxide myoglobin.
Authors: Rousseau D L , Argade P V
Published: May 2006

The photoproduct of carbon monoxide myoglobin generated at 4 K and lower has a resonance Raman spectrum characteristic of a high-spin heme but in which the high-frequency core size-sensitive lines are at lower frequency than those in the deoxy preparation. Such differences are not detected in the photoproduct generated at higher temperatures (50 K) or in that generated at room temperature with 10-nsec pulses. The data indicate that at the low temperature (4 K), the heme in the photoproduct is not fully relaxed, and from the data we conclude that the photoproduct has an expanded porphyrin core. We infer that the core size exceeds that in deoxymyoglobin because the rigid protein prevents the highspin iron atom from moving to its full out-of-plane displacement at the very low temperatures.
Complete sequence of HLA-B27 cDNA identified through the characterization of structural markers unique to the HLA-A, -B, and -C allelic series.
Authors: Szöts H , Riethmüller G , Weiss E , Meo T
Published: May 2006

Antigen HLA-B27 is a high-risk genetic factor with respect to a group of rheumatoid disorders, especially ankylosing spondylitis. A cDNA library was constructed from an autozygous B-cell line expressing HLA-B27, HLA-Cw1, and the previously cloned HLA-A2 antigen. Clones detected with an HLA probe were isolated and sorted into homology groups by differential hybridization and restriction maps. Nucleotide sequencing allowed the unambiguous assignment of cDNAs to HLA-A, -B, and -C loci. The HLA-B27 mRNA has the structural features and the codon variability typical of an HLA class I transcript but it specifies two uncommon amino acid replacements: a cysteine in position 67 and a serine in position 131. The latter substitution may have functional consequences, because it occurs in a conserved region and at a position invariably occupied by a species-specific arginine in humans and lysine in mice. The availability of the complete sequence of HLA-B27 and of the partial sequence of HLA-Cw1 allows the recognition of locus-specific sequence markers, particularly, but not exclusively, in the transmembrane and cytoplasmic domains.
Cloning of two genes that are specifically expressed in activated cytotoxic T lymphocytes.
Authors: Lobe C G , Havele C , Bleackley R C
Published: May 2006

The intracellular events leading to cytotoxic T-lymphocyte (CTL) activation and the molecular mechanism of target-cell lysis remain largely unknown. Differential hybridization analysis of a library constructed from a cloned CTL line was used to identify sequences specifically expressed in CTL. Two clones were selected for extensive analysis. No evidence for expression of their mRNAs was found in helper T-cell lines, thymocytes, lipopolysaccharide-activated B cells, interferon-treated natural killer cells, and a number of nonlymphoid cells. Blot-hybridization analysis of CTL mRNA revealed that one clone detected a single 900-nucleotide mRNA, whereas the other hybridized to two mRNAs of 900 and 1200 nucleotides, respectively. The maximum expression of these mRNAs precedes the peak of cytotoxicity in an in vitro allogeneic or mitogen-induced cytotoxic response by 24 hr; thus, they both fulfill the primary prerequisite for genes encoding proteins that are important in either CTL activation or in the lytic process itself.
Four virally determined nuclear antigens are expressed in Epstein-Barr virus-transformed cells.
Authors: Kallin B , Dillner J , Ernberg I , Ehlin-Henriksson B , Rosén A , Henle W , Henle G , Klein G
Published: May 2006

The expression of Epstein-Barr virus (EBV)-determined antigens associated with growth-transformation of B cells was studied by immunoblotting with human sera from healthy donors. Four antigens were detected in EBV-carrying cell lines and in B lymphocytes early after infection with the transforming B95-8 substrain of virus. They were not found in uninfected cells, nor could they be demonstrated with sera lacking antibodies to EBV antigens. All four antigens were nuclear. Each of them varied in size in the different cell lines. The two antigens with the lowest molecular weight were identified as EBV-determined nuclear antigens (EBNAs) 1 and 2. The two high molecular weight antigens (140-160 kDa and 150-180 kDa, respectively) were detected with 6 of 16 EBV antibody-positive sera. These proteins appeared to be antigenically unrelated to each other and to EBNAs 1 and 2 and were designated EBNAs 3 and 4. Like EBNAs 1 and 2, they bound to double- and single-stranded DNA in vitro.
Immunoreactivity for a calmodulin-dependent protein kinase is selectively increased in macaque striate cortex after monocular deprivation.
Authors: Hendry S H , Kennedy M B
Published: May 2006

Immunocytochemical methods were used to localize type II Ca2+/calmodulin-dependent protein kinase in the macaque primary visual cortex. Neurons that stain for the kinase include both pyramidal and nonpyramidal cells and they appear to form a subset of cortical neurons. They are densely packed in layers II and IVB, somewhat more sparse in layers III, IVC beta, and VI, and nearly absent in layer V. In normal animals the distribution of kinase-positive cells within each layer is relatively uniform. However, in animals in which one eye is removed 7-14 days before sacrifice or sutured shut for 9 or 11 weeks, the cells in layer IVC beta are divided into alternating lightly and darkly stained bands. Comparison of immunocytochemically stained sections with adjacent sections stained for the mitochondrial enzyme, cytochrome oxidase, reveals that the kinase staining increases in ocular dominance columns originally driven by the removed or closed eye. These findings suggest that either the concentration of type II Ca2+/calmodulin-dependent protein kinase or its accessibility to the antibody probe increases dramatically and selectively in neurons of macaque primary visual cortex that have been deprived of their normal visual input. This may indicate that changing levels of activity in cortical neurons can alter their regulatory machinery.
The nil Hecke ring and cohomology of
G/P
for a Kac-Moody group
G
Authors: Kostant Bertram , Kumar Shrawan
Published: May 2006

Let
G
be the group with Borel subgroup
B
, associated to a Kac-Moody Lie algebra [unk] (with Weyl group
W
and Cartan subalgebra [unk]). Then
H
*
(
G/B
) has, among others, four distinguished structures (
i
) an algebra structure, (
ii
) a distinguished basis, given by the Schubert cells, (
iii
) a module for
W
, and (
iv
) a module for Hecke-type operators

A
w

, for
w
[unk]
W
. We construct a ring
R
, which we refer to as the nil Hecke ring, which is very simply and explicitly defined as a functor of
W
together with the
W
-module [unk] alone and such that all these four structures on
H
*
(
G/B
) arise naturally from the ring
R
.
Benzyl acetates as attractants for the male oriental fruit fly,
Dacus dorsalis
, and the male melon fly,
Dacus cucurbitae
Authors: Metcalf Robert L. , Metcalf Esther R. , Mitchell W. C.
Published: May 2006

Fifty compounds related to benzyl acetate were evaluated quantitatively as attractants to the male oriental fruit fly (
Dacus dorsalis
) and the male melon fly (
Dacus cucurbitae
). Thienylmethyl acetate was nearly as attractive as benzyl acetate to both species, but cyclohexylmethyl acetate was completely unattractive, emphasizing the role of the planar aromatic ring in receptor interaction. Although benzyl acetate was equally attractive to both species,
para
substituents invariably reduced attraction to the oriental fruit fly. A number of derivatives, including
p
-hydroxy-,
p
-methoxy-,
p
-acetoxy-, and
p
-cyanobenzyl acetates were highly attractive to the melon fly. Selective fluorination indicated that the polarizability of the carbonyl carbon is important in receptor interaction with the melon fly but is not specifically involved in the oriental fruit fly. Attraction to the melon fly was lost with
ortho, meta
, or dimethoxy substitution of benzyl acetate, but the 3,4- and 2,5-dimethoxybenzyl acetates were attractive to the oriental fruit fly. These results are interpreted in terms of specific olfactory receptor interactions for the two species of fruit flies.
Large introns in the 3' end of the gene for the pro alpha 1 (IV) chain of human basement membrane collagen.
Authors: Soininen R , Tikka L , Chow L , Pihlajaniemi T , Kurkinen M , Prockop D J , Boyd C D , Tryggvason K
Published: May 2006

Using a recently characterized cDNA clone (HT-21) coding for the pro alpha 1 (IV) chain of human type IV procollagen, we have isolated three clones from a bacterio-phage lambda Charon 4A library of human genomic DNA. The intron/exon structure of the pro alpha 1 (IV) genomic clones was analyzed by heteroduplex electron microscopy and nucleotide sequencing. The analysis showed that the introns separating exons 2-9 are large and have a total length of over 12,000 base pairs (bp). Six of seven exons at the 3' end of the gene coded for -Gly-Xaa-Yaa-repeats of the collagenous part of the chain. Five of the -Gly-Xaa-Yaa- coding exons (numbers 5-9) varied in size between 72 bp and 134 bp, and none of them were 54 bp or multiples thereof. A sixth exon (exon 4) was a junction exon containing 71 bp coding for -Gly-Xaa-Yaa- sequences and 142 bp coding for the carboxyl-terminal noncollagenous domain (NC-1). The seventh exon (exon 3, 178 bp) coded for sequences of the NC-1 domain. Five of the six -Gly-Xaa-Yaa- coding exons began with the second base coding for glycine, and only one exon began with a complete glycine codon at the 5' end. The results (i) suggest that the gene for the pro alpha 1(IV) chain of human basement membrane collagen is significantly larger than the genes for fibrillar collagens and (ii) show that it lacks the 54-bp exon repeats characteristic of fibrillar collagen genes.
Bovine galactosyltransferase: identification of a clone by direct immunological screening of a cDNA expression library.
Authors: Shaper N L , Shaper J H , Meuth J L , Fox J L , Chang H , Kirsch I R , Hollis G F
Published: May 2006

A 1.3-kilobase cDNA clone (7A) coding for bovine galactosyltransferase (glycoprotein 4-beta-galactosyltransferase, EC 2.4.1.38) was isolated from a lambda gt11 expression library by immunological screening with monospecific polyclonal antisera to the affinity-purified bovine enzyme. The nucleotide sequence of this clone predicts an open reading frame that starts at the 5' end of the insert and codes for a polypeptide of 334 amino acids with Mr 37,645. Based on a Mr of 57,000 for the membrane-bound enzyme this clone accounts for approximately 61% of the coding sequence. Portions of the predicted amino acid sequence matched the six tryptic peptides isolated from affinity-purified bovine galactosyltransferase. Clone 7A hybridizes to a 4.8-kilobase bovine mRNA and identifies multiple EcoRI restriction fragments in bovine, murine, and human DNA.
Enhanced c-Ki-ras expression associated with Friend virus integration in a bone marrow-derived mouse cell line.
Authors: George D L , Glick B , Trusko S , Freeman N
Published: May 2006

We have investigated the molecular basis for a 25- to 30-fold overexpression of the c-Ki-ras oncogene in a mouse bone marrow-derived, early myeloid cell line, 416B. Southern blot hybridizations revealed that the 416B cells contain a rearranged c-Ki-ras gene in addition to an apparently normal gene. Molecular cloning and DNA sequence analyses demonstrated that the rearrangement involves the insertion of a 3.5-kilobase-pair segment of Friend virus that includes the envelope gene (env) and 3' long terminal repeat. The Friend provirus is positioned between a 5' nontranslated exon (exon phi) and the first coding exon (exon 1) of the c-Ki-ras gene in the same transcriptional orientation. Results of RNA blot analyses indicate that transcription from the rearranged gene initiates at a promoter that excludes sequences in exon phi. The data support the hypothesis that enhanced c-Ki-ras expression in the 416B cells results from integration of a Friend provirus within this gene.
Identification of a common nucleotide sequence in the 3'-untranslated region of mRNA molecules specifying inflammatory mediators.
Authors: Caput D , Beutler B , Hartog K , Thayer R , Brown-Shimer S , Cerami A
Published: May 2006

Recently, cDNA sequences have been reported for both human and murine tumor necrosis factor (TNF; cachectin). The coding region of the TNF genes is highly conserved between man and mouse; 80% homology is apparent at the amino acid level. We now observe that a 33-nucleotide sequence, comprised entirely of A and T residues and located in the 3'-untranslated region, is conserved in toto in the murine and human TNF mRNAs. Since the 3'-untranslated region is normally not conserved, we reasoned that this sequence might play a regulatory role. We identified a consensus sequence (TTATTTAT) present in the 3'-untranslated region of both human and mouse TNF mRNAs, as well as the mRNAs encoding human lymphotoxin, human colony stimulating factor, human and mouse interleukin 1, human and rat fibronectin, and most of the sequenced human and mouse interferons. All of these mRNAs, except the lymphotoxin mRNA, lack homology to the TNF mRNAs in the coding region. The consensus sequence is uncommon among mammalian mRNAs in general, but it appears with a frequency greater than chance alone would dictate, suggesting that it may serve a specific regulatory function among the mRNAs in which it is found. It is particularly prevalent among mRNAs encoding proteins related to the inflammatory response.
Primary electron transfer reactions in modified reaction centers from
Rhodopseudomonas sphaeroides
Authors: Shuvalov V. A. , Duysens L. N. M.
Published: May 2006

Absorption spectra were measured by means of an optical multichannel analyzer in
Rhodopseudomonas sphaeroides
R-26 reaction centers (RCs) modified by treatment with NaBH
4
at various times (≥1 ps) after the onset of a short excitation flash at 880 nm. Most of these RCs (75-95%) have only one “monomeric” bacteriochlorophyll-800 (B
1
) molecule and are as active as the original RCs. The duration of the excitation and measuring pulses was ≈33 ps. If the center of the excitation pulse preceded the center of the measuring pulse by 36-40 ps, the formation of a state P
E
(early state), which is converted to the state P
F
(P
+
bacteriopheophytin
-
) in 4 ± 1 ps (1/
e
time), was observed. Also the kinetics and the spectrum of the stimulated emission (reflecting the kinetics and the emission spectrum of the excited state P
*
) were determined. The difference spectrum of the state P
E
approximately equals the sum of the spectra of the states P
*
(≈65%) and
1
[P
+
B
1
-
] (≈35%). This indicates that B
1
-
is an intermediate in the electron transfer from P
*
to bacteriopheophytin, H
1
, transferring this electron with a rate constant of (4 × 0.35 ps)
-1
= 7 × 10
11
s
-1
.
Quantitation of insertion sequence IS10 transposase gene expression by a method generally applicable to any rarely expressed gene.
Authors: Raleigh E A , Kleckner N
Published: May 2006

We have found that IS10 transposase is synthesized in tiny amounts, about 0.15 polypeptide chain per cell per generation on average, as judged from the beta-galactosidase activity of a single chromosomal copy of a suitable transposase-lacZ gene fusion. Enzymatic activity from the fusion gene is a factor of 10 lower in a permeabilized whole cell assay than in cell extracts. Probably, most cells contain fewer than four polypeptide chains, and these chains can assemble into active tetramers only after cell disruption. This interpretation permits formulation of two equations relating enzyme activities to transcription and translation rates, solution of which reveals that the fusion gene is expressed at the average rate of only 0.25 transcript per cell per generation, with an average of only 0.58 translation product per transcript. This methodology is generally applicable to analysis of any gene from which fewer than four polypeptide chains are synthesized per cell per generation.
Somatic evolution of variable region structures during an immune response.
Authors: Wysocki L , Manser T , Gefter M L
Published: May 2006

Immunization of strain A mice with p-azophenylarsonate-conjugated protein stimulates B cells that synthesize anti-p-azophenylarsonate antibodies. A large fraction of these cells produce antibodies with variable (V) regions encoded by a single heavy chain V gene segment together with multiple combinations of diversity, heavy chain joining, light chain variable, and light chain joining gene segments. Early in the immune response, these V regions are not somatically mutated. One of these V regions is initially expressed by only a minority of the responding B cells but binds p-azophenylarsonate with the highest affinity. After a secondary immunization, B cells synthesizing mutated derivatives of this single V region dominate the response and bind p-azophenylarsonate with even higher affinity than does the unmutated V region. These results suggest that antigen directs both the expression of the immune repertoire and the amplification of V region diversity by a sequential process of clonal selection of B cells expressing receptor antibodies encoded by unmutated V genes, induction of mutation in the V genes expressed by the selected cells, and reselection of B cells expressing antibodies with mutated V regions of higher affinity.
Resistance to human respiratory syncytial virus (RSV) infection induced by immunization of cotton rats with a recombinant vaccinia virus expressing the RSV G glycoprotein.
Authors: Elango N , Prince G A , Murphy B R , Venkatesan S , Chanock R M , Moss B
Published: May 2006

A cDNA copy of the G glycoprotein gene of human respiratory syncytial virus (RSV) was placed under control of a vaccinia virus promoter and inserted into the thymidine kinase locus of the vaccinia virus genome. The recombinant vaccinia virus retained infectivity and expressed a 93-kDa protein that migrated with the authentic RSV G glycoprotein upon polyacrylamide gel electrophoresis. Glycosylation of the expressed protein and transport to the cell surface were demonstrated in the absence of other RSV proteins. Cotton rats that were inoculated intradermally with the infectious recombinant virus produced serum antibody to the G glycoprotein that neutralized RSV in vitro. Furthermore, the vaccinated animals were resistant to lower respiratory tract infection upon intranasal inoculation with RSV and had reduced titers of RSV in the nose.
Solution of an inverse problem for fractals and other sets
Authors: Barnsley M. F. , Ervin V. , Hardin D. , Lancaster J.
Published: May 2006

The problem of providing succinct approximate descriptions of given bounded subsets of R

n

can be solved by application of the contraction mapping principle.
Glycosylation of ovalbumin in a heterologous cell: analysis of oligosaccharide chains of the cloned glycoprotein in mouse L cells.
Authors: Sheares B T , Robbins P W
Published: May 2006

In an effort to understand the factors that influence protein glycosylation, we are studying the expression of the chicken ovalbumin gene in a heterologous cell. Ovalbumin synthesized in mouse L-353 cells is glycosylated, as judged by incorporation of [3H]mannose and susceptibility to endo-beta-N-acetylglucosaminidases. Sequential digestion of ovalbumin synthesized in L cells with trypsin and chymotrypsin yields material migrating as one peak on HPLC coincident with similarly treated material from chicken ovalbumin, suggesting that the protein synthesized in L cells is glycosylated at the correct site. As in the case of chicken ovalbumin, the protein synthesized in L cells contains large amounts of hybrid oligosaccharides. Approximately 50% of the [3H]mannose incorporated into ovalbumin secreted by L cells is found in such hybrid structures. These results suggest that it is the polypeptide chain of ovalbumin that is responsible for proper glycosylation and subsequent processing of a substantial fraction of the oligosaccharide chains to hybrid structures. However, differences do exist between ovalbumin synthesized in L cells and the chicken glycoprotein. The hybrid oligosaccharides of ovalbumin secreted by L cells are completely sialylated and do not contain a bisecting GlcNAc residue, distinguishing them from hybrid chains in chicken ovalbumin. In addition to high-mannose and hybrid oligosaccharide chains, ovalbumin synthesized in L cells contains oligosaccharides of the complex type. To date, this type of sugar chain has not been observed in chicken ovalbumin. These differences in fine structure, between the oligosaccharides derived from ovalbumin secreted by L cells and those known to be present in the chicken egg glycoprotein, suggest that the cell type also plays a role in oligosaccharide processing.
Visualization of the paranemic joining of homologous DNA molecules catalyzed by the RecA protein of Escherichia coli.
Authors: Christiansen G , Griffith J
Published: May 2006

In reactions catalyzed by the RecA protein of Escherichia coli, synapsis between two DNA molecules is believed to occur even in the absence of free homologous DNA ends and to involve a metastable interaction termed paranemic joining. We have used electron microscopic methods to visualize synapse formation between supertwisted M13 double-stranded DNA (dsDNA) and linear M13 mp7 single-stranded DNA (ssDNA) with non-M13 sequences at its ends. These non-M13 sequences block strand invasion and make this pairing equivalent to the joining of two fully circular molecules. We observed a high frequency of joining when the ssDNA was initially assembled into presynaptic filaments with RecA protein. Cleavage of the dsDNA in the joined complexes by Hpa I revealed that the joint was at a site of homology. In these joints, the dsDNA entered the presynaptic filament over a length of 360 +/- 80 base pairs, not visibly altering its ultrastructure, and then dissociated from the filament. Although the dsDNA in the complexes appeared topologically relaxed, deproteinization released supertwisted dsDNA, indicating that the dsDNA was unwound by 34 degrees per base pair in the paranemic joint. When supertwisted M13 dsDNA was paired with circular M13 ssDNA, similar joints were observed and both DNA circles appeared topologically relaxed.
Human recombinant transforming growth factor alpha stimulates bone resorption and inhibits formation in vitro.
Authors: Ibbotson K J , Harrod J , Gowen M , D'Souza S , Smith D D , Winkler M E , Derynck R , Mundy G R
Published: May 2006

Human recombinant transforming growth factor alpha (TGF alpha), which binds to the epidermal growth factor (EGF) receptor and causes several biological effects similar to those caused by EGF, was compared with murine EGF for its effects on a number of parameters of bone cell metabolism. TGF alpha stimulated bone resorption in two organ culture systems, the fetal rat long bone and neonatal mouse calvarial systems. TGF alpha stimulated bone resorption at concentrations as low as 0.1 ng/ml. TGF alpha effects on bone resorption in mouse calvariae were inhibited by indomethacin, suggesting that, like EGF, its effects were mediated by prostaglandin synthesis. TGF alpha had a different time course of action on bone resorption from that of EGF, causing more rapid release of previously incorporated 45Ca from bone cultures, suggesting that TGF alpha does not function on bone as a simple EGF analogue. TGF alpha also caused effects on osteoblast function resembling those of EGF. It inhibited alkaline phosphatase activity in cultured rat osteosarcoma cells with the osteoblast phenotype and inhibited collagen synthesis in fetal rat calvaria at concentrations of 1.0 ng/ml. The lowest concentration of TGF alpha (expressed as nanogram equivalents of EGF per ml) required to produce a response in all of the systems tested was about 1/10th of that needed for EGF to produce a similar effect. These results indicate that TGF alpha is a potent stimulator of bone resorption and inhibitor of bone formation as assessed by inhibition of collagen synthesis and alkaline phosphatase activity and are consistent with the hypothesis that TGF alpha may be responsible, at least in part, for the bone resorption associated with some tumors.
Separation and properties of cellular and scrapie prion proteins.
Authors: Meyer R K , McKinley M P , Bowman K A , Braunfeld M B , Barry R A , Prusiner S B
Published: May 2006

Purified preparations of scrapie prions contain a sialoglycoprotein of Mr 27,000-30,000, designated PrP 27-30, which is derived from the scrapie prion protein [Mr, 33,000-35,000 (PrP 33-35Sc)] by limited proteolysis. Under these same conditions of proteolysis, a cellular protein of the same size (PrP 33-35C) is completely degraded. Subcellular fractionation of hamster brain showed that both PrP 33-35Sc and PrP 33-35C were found only in membrane fractions. NaCl, EDTA, and osmotic shock failed to release the prion proteins from microsomal membranes. Electron microscopy of these microsomal fractions showed membrane vesicles but not prion amyloid rods. Detergent treatment of scrapie-infected membranes solubilized PrP 33-35C, while PrP 33-35Sc aggregated into amyloid rods; the concentration of PrP 33-35C was similar to that recovered from analogous fractions prepared from uninfected control brains. The apparent amphipathic character of the PrP 33-35Sc may explain the association of scrapie infectivity with both membranes and amyloid filaments.
A molecular mechanism for sensory adaptation based on ligand-induced receptor modification.
Authors: Knox B E , Devreotes P N , Goldbeter A , Segel L A
Published: May 2006

Physiological responses mediated by cell-surface receptors frequently adapt or "desensitize" (i.e., terminate despite persistent occupancy of receptors by ligand). Binding of ligands to the external domains of a wide variety of surface receptors induces covalent modification of their cytoplasmic domains. A mechanism is presented in which the variety of receptor states generated by ligand binding and covalent modification act together to regulate physiological responsiveness. The development of the model is guided by observations of adaptation for chemotaxis in Escherichia coli and adenylate cyclase activation in Dictyostelium. The general features of the marked response and eventual exact adaptation predicted by the model match those observed in the experimental systems.
Developmental regulation of two genes encoding ribulose-bisphosphate carboxylase small subunit in pea and transgenic petunia plants: Phytochrome response and blue-light induction
Authors: Fluhr Robert , Chua Nam-Hai
Published: May 2006

We describe the effects of light quality on the light-induced expression of two genes (
rbcS-3A
and
rbcS-3C
) encoding the ribulose-1,5-bisphosphate carboxylase small subunit of pea plants. These two genes exhibit distinctive photoresponses depending on the developmental stages of the leaves. In etiolated primary leaves, changes in the
rbcS-3A
and
rbcS-3C
transcript levels are regulated by phytochrome. In mature green leaves, on the other hand, the transcript levels are modulated by a putative blue-light photoreceptor working in concert with phytochrome. We transferred the two pea
rbcS
genes into the petunia genome and found that their phytochrome (
rbcS-3A
and -
3C
) and blue-light responses (
rbcS-3C
) are recapitulated in the appropriate developmental stages of the transgenic plants. Moreover, the transgenic plants express the pea
rbcS
genes in leaves at levels close to those in pea leaves. Our results show that 0.4 kilobases of the
rbcS-3A
and 2 kilobases of the
rbcS-3C
upstream regions are sufficient for these photoresponses.
Oxidation as a possible mechanism of cellular aging: vitamin E deficiency causes premature aging and IgG binding to erythrocytes.
Authors: Kay M M , Bosman G J , Shapiro S S , Bendich A , Bassel P S
Published: May 2006

Senescent-cell antigen is a "neo-antigen" that appears on the surface of senescent cells and initiates IgG binding and cellular removal. As an approach to evaluating oxidation as a possible mechanism for generation of senescent-cell antigen, we studied erythrocytes from vitamin E-deficient rats. Vitamin E is localized primarily in cellular membranes. Its major role is the termination of free-radical chain reactions propagated by the polyunsaturated fatty acids of membrane propagated by the polyunsaturated fatty acids of membrane phospholipids. Results of our studies indicate that erythrocytes of all ages from vitamin E-deficient rats behave like old erythrocytes from normal rats, as determined by their susceptibility to phagocytosis, IgG binding, anion transport ability, and glyceraldehyde-3-phosphate dehydrogenase activity. Increased breakdown products of band 3 were observed with immunoblotting in membranes of erythrocytes from vitamin E-deficient rats. Breakdown products of band 3 are known to increase as cells age in normal individuals. The data suggest that oxidation may be a possible mechanism for erythrocyte aging and generation of senescent-cell antigen in vivo.
cAMP increases junctional conductance and stimulates phosphorylation of the 27-kDa principal gap junction polypeptide.
Authors: Saez J C , Spray D C , Nairn A C , Hertzberg E , Greengard P , Bennett M V
Published: May 2006

Membrane-permeant cAMP derivatives (dibutyryl- and 8-bromo-cAMP) increase gap-junctional conductance within minutes when applied to voltage-clamped pairs of rat hepatocytes. Glucagon also increases junctional conductances, but the response has a more rapid onset and is more rapidly reversible. The glucagon effect can be prevented by intracellular injection of the protein inhibitor of the cAMP-dependent protein kinase (Walsh inhibitor), indicating that the catalytic subunit of cAMP-dependent protein kinase is directly involved. The 27-kDa major gap junction polypeptide is phosphorylated when liver cells dissociated into small groups are incubated with 32P. Addition of 8-bromo-cAMP to cells increases the incorporation of 32P into the 27-kDa junctional protein. Serine is the amino acid residue that is phosphorylated. When isolated liver gap junctions are incubated in the presence of catalytic subunit of the cAMP-dependent protein kinase, the 27-kDa gap junction polypeptide is phosphorylated with low stoichiometry on serine. The rapid increases in gap junctional conductance caused by agents that elevate cAMP and phosphorylation of the gap junction protein by cAMP-dependent protein kinase suggest that cAMP-dependent phosphorylation of the gap junction channel modulates the conductance of liver gap junctions.
Involvement of Lyt-2 and L3T4 in activation of hapten-specific Lyt-2+ L3T4+ T-cell clones.
Authors: Fazekas de St Groth B , Gallagher P F , Miller J F
Published: May 2006

The murine T-cell surface molecules Lyt-2 and L3T4 play a role in the activation of antigen-specific T cells. The currently accepted model for the function of these molecules proposes that Lyt-2 and L3T4 increase the overall avidity of the interaction between the T-cell antigen receptor and antigen in association with the major histocompatibility complex (MHC) molecules on the antigen-presenting cell. We have used two unusual Lyt-2+ L3T4+ class II MHC-restricted T-cell clones to test whether Lyt-2 can substitute for L3T4 when the T-cell antigen receptor is class II MHC-restricted. Monoclonal antibodies against L3T4 profoundly inhibited antigen-induced lymphokine production by both T-cell clones. Anti-Lyt-2 monoclonal antibody had no effect. These results strongly suggest that L3T4 and the class II-restricted T-cell antigen receptors are physically close during antigen recognition, probably as part of a multimolecular complex from which Lyt-2 is excluded. The ability of L3T4 but not Lyt-2 to participate in such a complex with class II-restricted T-cell antigen receptors may explain the striking correlation between class II restriction and L3T4 expression in the peripheral T-cell pool.
Site-directed mutagenesis of an invariant amino acid residue at the variable-diversity segments junction of an antibody.
Authors: Sharon J , Gefter M L , Manser T , Ptashne M
Published: May 2006

Structural analysis of 21 murine A/J antibodies specific for the hapten p-azobenzenearsonate (Ars), and bearing the major cross reactive idiotype (IdCRI), has revealed an invariant amino acid residue, serine, encoded by the variable-diversity gene segments junction of the heavy chain. To test whether this serine residue is essential for Ars binding, we changed it either to alanine or to threonine by oligonucleotide-directed mutagenesis of a heavy chain gene. Genes containing the mutations were separately introduced into mouse hybridoma cells producing the homologous light chain, and the resulting proteins were tested for antigen binding and idiotypic expression. Whereas the serine to threonine mutant retains full antigen binding activity, the serine to alanine mutant does not bind either to Ars-bovine serum albumin-Sepharose or to the Ars-tyrosine hapten. Both mutants show the same reactivity as wild type towards a series of anti-idiotypic antibodies. These results suggest that a hydroxyl group at the variable-diversity gene segments junction of A/J anti-Ars antibodies is essential for antigen binding.
Amiloride-sensitive Na channels from the apical membrane of the rat cortical collecting tubule.
Authors: Palmer L G , Frindt G
Published: May 2006

Currents through individual Na channels in the apical membrane of the rat cortical collecting tubule were resolved by using the patch-clamp technique. In cell-attached patches, the channels had a conductance of 5 pS with 140 mM NaCl in the pipet. The conductance was a saturable function of external Na, with a maximal value of about 8 pS and a half saturation at about 75 mM Na. In excised inside-out patches, the selectivity of the channels for Na over K was estimated from reversal potentials to be at least 10:1. The channels underwent spontaneous transitions between open and closed states. Both states had mean lifetimes of 3-4 sec. Amiloride (0.5 microM) added to the pipet induced more frequent closures and openings of the channels and a reduction in the mean open time. These channels are presumed to mediate Na reabsorption by this nephron segment in vivo.
Temporal order of replication of Xenopus laevis 5S ribosomal RNA genes in somatic cells.
Authors: Gilbert D M
Published: May 2006

The timing of replication of both the somatic and oocyte-type 5S ribosomal RNA genes of Xenopus laevis was determined in cultured cells by using 5-bromodeoxyuridine labeling of DNA coupled with a retroactive synchrony technique employing the fluorescence-activated cell sorter (FACS). The somatic genes replicate very early in S phase, while the oocyte genes replicate very late. These experiments provide direct evidence for a model [Gottesfeld, J. & Bloomer, L. S. (1982) Cell 28, 781-791] in which the transcription-activating factor TFIIIA is required at the time of Xenopus 5S rRNA gene replication to facilitate transcription of these genes.
A translocated human c-myc oncogene is altered in a conserved coding sequence.
Authors: Murphy W , Sarid J , Taub R , Vasicek T , Battey J , Lenoir G , Leder P
Published: May 2006

We have cloned and characterized a c-myc (now designated MYC) oncogene that had been translocated into the mu switch region of the immunoglobulin heavy chain locus in a Burkitt lymphoma cell line. The breakpoint of the translocation occurs within the first intron of the c-myc gene, thereby separating the untranslocated first exon from the two coding exons. Transcription from the translocated gene arises from a cryptic promoter within the first intron, which produces a 438-nucleotide untranslated 5' region. The amino acid sequence of the protein encoded by the c-myc gene has been substantially altered. In particular, a compensating set of frameshift mutations alters a string of 24 amino acids in a region of the protein tightly conserved in human, mouse, and chicken c-myc genes as well as in the human N-myc and L-myc oncogenes. Despite this, the mutated gene retains a reduced transforming ability in a rat embryo fibroblast focus-formation assay.
Gene encoding T-cell-activating protein TAP maps to the Ly-6 locus.
Authors: Reiser H , Yeh E T , Gramm C F , Benacerraf B , Rock K L
Published: May 2006

Recently we described two murine T-cell membrane proteins, TAP (T-cell-activating protein) and TAPa (TAP-associated protein). Previous experiments suggested that TAP is involved in physiologic T-cell activation. The subject of this report is a genetic analysis of these molecules. TAP and TAPa map to the Ly-6 locus. The relationship of these molecules to other antigens encoded in this locus is examined. Based on tissue distribution, molecular structure, and functional properties, TAP is distinct from any previously described Ly-6 antigen, whereas TAPa is probably identical to the 34-11-3 antigen. TAP and TAPa are coexpressed on all cell types examined so far. Moreover, comparative studies demonstrate a complex developmentally regulated pattern in the expression of molecules encoded in this locus.
Specificity analysis of human monoclonal antibodies reactive with cell surface and intracellular antigens.
Authors: Cote R J , Morrissey D M , Houghton A N , Thomson T M , Daly M E , Oettgen H F , Old L J
Published: May 2006

Over 4350 human Ig-secreting hybrids have been generated through the fusion of human lymphocytes with NS-1 (mouse), LICR-2, SKO-007, GM4672, or UC729-6 (human) myeloma and lymphoblastoid cell lines. NS-1 proved to be the most satisfactory fusion partner, and 83% of the stable Ig-secreting clones were derived from NS-1 fusions. Three hundred five hybrids produced human monoclonal antibodies (hmAb) reactive with cell surface or intracellular antigens expressed by cultured human tumor cell lines, and 111 of these have undergone detailed serological specificity analysis. Several general points have emerged from our study of hmAb: A significant proportion of the human B-cell clones produce antibody reactive with cellular antigens. The majority of these antigens have an intracellular location and are broadly distributed. Intracellular and cell surface differentiation antigens and other antigens with restricted distribution have been defined by hmAb, including two cell surface antigens not detected on normal cells. The relationship of these findings to cancer is unclear, as hmAb reactive with antigens showing distinctive distribution have been generated from the lymphocytes of normal individuals as well as tumor-bearing patients.
Chromosomal breakpoints and structural alterations of the c-myc locus differ in endemic and sporadic forms of Burkitt lymphoma.
Authors: Pelicci P G , Knowles D M , Magrath I , Dalla-Favera R
Published: May 2006

We have examined the position of the chromosomal breakpoint relative to the human c-myc gene (MYC) and the presence of other structural alterations of the same locus in 19 fresh samples of Burkitt lymphoma (BL) and 13 BL-derived cell lines. This panel includes the two pathogenetic forms of BL: the endemic (African-type) BL (eBL) and sporadic (American-type) BL (sBL). In all cases tested, including fresh samples and cell lines, structural alterations of the 5' portion of the gene were detected, suggesting that they may be necessary for c-myc activation. However, the site of chromosomal breakpoint and the type of structural alterations differ in eBL and sBL. In 16 of 18 sBL cases, chromosomal translocation truncates the gene within a 5' region that includes the first intron, the first exon, and 5' flanking sequences. Conversely, in 14 of 14 eBL samples, the chromosomal breakpoint is located outside the c-myc locus, yet the same 5' sequences are affected by several mutations identifiable by restriction enzyme polymorphisms. Different genetic mechanisms may therefore be involved in chromosomal translocation/c-myc activation, and these differences may be a function of differences in the stage of differentiation of eBL versus sBL.