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PubMed1994

The functions and consensus motifs of nine types of peptide segments that form different types of nucleotide-binding sites.

From an analysis of current data on 16 protein structures with defined nucleotide-binding sites consensus motifs were determined for the peptide segments that form such nucleotide-binding sites. This was done by using the actual residues shown to contact ligands in the different protein structures, plus an additional 50 sequences for various kinases. Three peptide segments are commonly required to form the binding site for ATP or GTP. Binding motif Kinase-1a is found in almost all sequences examined, and functions in binding the phosphates of the ligand. Variant versions, comparable to Kinase-1a, are found in a subset of proteins and appear to be related to unique functions of those enzymes. Motif Kinase-2 contains the conserved aspartate that coordinates the metal ion on Mg-ATP. Motif Kinase-3 occurs in at least four versions, and functions in binding the purine base or the pentose. Two protein structures show ATP-binding at a separate regulatory site, formed by the motifs Regulatory-1 and Regulatory-2. Structures for adenylate kinase and guanylate kinase show three different sequence motifs that form the binding site for a nucleoside monophosphate (NMP). NMP-1 and NMP-2 bind to the pentose and phosphate of the bound ligand. NMP-1 is found in almost all the kinases that phosphorylate AMP, CMP, GMP, dTMP, or UMP. NMP-3a is found in kinases for AMP, GMP, and UMP, while NMP-3b binds only GMP. For the binding of NTPs, three distinct types of nucleotide-binding fold structures have been described. Each structure is associated with a particular function (e.g. transfer of the gamma-phosphate, or of the adenylate to an acceptor) and also with a particular spatial arrangement of the three Kinase segments evident in the linear sequence for the protein.

Adenosine Triphosphate↗

PubMed2003

SNARE selectivity of the COPII coat.

The COPII coat buds transport vesicles from the endoplasmic reticulum that incorporate cargo and SNARE molecules. Here, we show that recognition of the ER-Golgi SNAREs Bet1, Sed5, and Sec22 occurs through three binding sites on the Sec23/24 subcomplex of yeast COPII. The A site binds to the YNNSNPF motif of Sed5. The B site binds to Lxx-L/M-E sequences present in both the Bet1 and Sed5 molecules, as well as to the DxE cargo-sorting signal. A third, spatially distinct site binds to Sec22. COPII selects the free v-SNARE form of Bet1 because the LxxLE sequence is sequestered in the four-helix bundle of the v-/t-SNARE complex. COPII favors Sed5 within the Sed5/Bos1/Sec22 t-SNARE complex because t-SNARE assembly removes autoinhibitory contacts to expose the YNNSNPF motif. The COPII coat seems to be a specific conductor of the fusogenic forms of these SNAREs, suggesting how vesicle fusion specificity may be programmed during budding.

Amino Acid Motifs↗

PubMed1988

Spatially deranged though temporally correct expression of Strongylocentrotus purpuratus actin gene fusion in transgenic embryos of a different sea urchin family.

We report the unexpected observation that cis-regulatory sequences of a Strongylocentrotus purpuratus actin gene, which direct a particular, lineage-specific pattern of embryonic expression, confer a completely different spatial pattern of expression when introduced into embryos of another sea urchin species. We utilized a fusion gene construct in which the bacterial chloramphenicol acetyl transferase (CAT) reporter gene is driven by CyIIIa actin regulatory sequences. We previously showed that the regulatory region that is included suffices to promote the accumulation of CAT mRNA in transgenic S. purpuratus embryos, on the same developmental schedule and in the same embryonic region, the aboral ectoderm, in which the CyIIIa actin gene is normally expressed (Flytzanis et al. 1987; Hough-Evans et al. 1987). When injected into zygotes of Lytechinus variegatus, which belongs to a different echinoid family, the expected temporal pattern of expression of CAT enzyme was observed. Thus, in both S. purpuratus and L. variegatus embryos, expression is activated at the early blastula stage, although this stage is attained several hours sooner in L. variegatus embryo cultures. Similar kinetics of CAT enzyme accumulation were obtained whether the gene was introduced directly into the L. variegatus zygote nucleus or into the cytoplasm. However, when examined by in situ hybridization, the transgenic L. variegatus embryos were found to display a totally new pattern of CAT mRNA accumulation. Copious CAT transcripts were detected not only in aboral ectoderm cells, but also in skeletogenic mesenchyme cells, gut cells, and oral ectoderm, all cell types that in the transgenic S. purpuratus controls are invariably devoid of detectable CAT transcripts.

Acetyltransferases↗

PubMed2007

Patterns of dispersal in Sumatran siamangs (Symphalangus syndactylus): preliminary mtDNA evidence suggests more frequent male than female dispersal to adjacent groups.

Gibbons of both sexes have been observed emigrating from their natal groups, but the consequences of dispersal in gibbons are poorly understood, and it is unclear whether these are the same for both sexes. I sequenced a 350-bp fragment of mitochondrial DNA from 18 adults in seven siamang (Symphalangus syndactylus) groups at the Way Canguk Research Station in southern Sumatra to assess patterns of matrilineal relatedness among and within siamang groups, and to assess their fit with different patterns of sex-specific dispersal. A total of 11 haplotypes were identified in the seven study groups; 50% of adult males in five contiguous groups shared a haplotype with a member of an immediately adjacent group, whereas only 16.7% of females shared a haplotype with a neighbor. The apparent difference persisted if only same-sex individuals were considered (37.5% of males vs. 0% of females). Four of the seven study groups contained two adult males and a single adult female. In three multimale groups, the three adults all had different haplotypes, suggesting that neither male was the retained adult offspring of the female, whereas in the fourth group, the haplotype of one male was identical with that of the adult female. The high diversity of haplotypes and the absence of clustering among female haplotypes in the study neighborhood suggest that female dispersal to territories adjacent to the natal group may be relatively rare. The presence of some clustering of male haplotypes suggests that shorter dispersal distances may be more common in males.

Animals↗

PubMed2005

Regulation of chromatin structure during thymic T cell development.

Development is the process whereby a multipotent cell gives rise, through series of divisions, to progeny with successively restricted potentials. During T cell development, the process begins with a multipotent hematopoietic stem cell (HSC) in the bone marrow, moves to the thymus where early T cells or thymocytes pass through signal-initiated developmental checkpoints, and ends in the periphery where mature T cells reside. At each step along this developmental pathway, T lymphocyte progenitors must be able to turn genes on and off, creating a specialized program of gene expression, to allow further development. How is gene expression coordinated? This review will summarize what has been learned about the function of chromatin structure in generating a "blueprint" of gene expression during T cell development. This will include discussion of mechanisms of chromatin remodeling, histone modification, and heritable gene silencing. In many cases, these processes are carried out by multi-protein complexes whose components are largely ubiquitously expressed. The spatial and temporal specificity of these complexes is contributed by sequence specific DNA binding factors, some of which are cell type restricted in their expression. This review will summarize research underway to identify these key genetic "targeters." Taken together, the research reviewed here provides a glimpse into the importance of regulation of chromatin structure in T cell development and the "players" involved.

Animals↗

PubMed2006

Strengths and weaknesses of dark-lumen MR colonography: clinical relevance of polyps smaller than 5 mm in diameter at the moment of their detection.

PURPOSE: To assess the clinical relevance of dark-lumen MR colonography (MRC) for the detection of colorectal lesions using conventional colonoscopy (CC) and histopathologic examinations as reference standard. MATERIALS AND METHODS: A total of 72 patients underwent MRC and CC. MRC was performed using a contrast-enhanced high spatial resolution T1 weighted 3D volumetric interpolated breathhold examination (VIBE)-sequence. All removed colorectal lesions were evaluated by an experienced pathologist. RESULTS: CC confirmed 65 polyps less than 5 mm in diameter. Non of those lesions could be detected using MRC. Just two (4%) of the 49 removed lesions smaller than 5 mm showed signs of dysplasia. Additionally, CC confirmed 25 polyps between 6-15 mm in diameter (MRC 22). All those 25 lesions were removed in CC. Only four (16%) of those polyps showed signs of dysplasia and malignancy (11, 13, 13 and 15 mm). CONCLUSION: Dark-lumen MRC failed to detect all polyps smaller than 5 mm in diameter which are generally not clinically relevant at the moment of their detection and thus can be kept under surveillance. However, MRC as a non-invasive imaging modality is a promising alternative to CC in the detection of clinically relevant polyps larger than 5 mm in diameter.

Adult↗

PubMed1988

In vivo localized proton spectroscopic studies of human gastrocnemius muscle.

In vivo proton magnetic resonance spectroscopy studies of gastrocnemius muscle were performed in six normal volunteers. Both spatially resolved spectroscopy (SPARS) and stimulated echo acquisition mode (STEAM) sequences were used for volume localization. A number of water suppression sequences have been combined with these localization schemes. Among the various techniques investigated in these studies, STEAM with an inversion pulse (T1-discriminated spectrum) seems to have the best potential for in vivo localized high-resolution proton spectroscopy studies of human muscle.

Adult↗

PubMed1991

Multiple-readout selective inversion recovery angiography.

We have developed a variation of selective inversion recovery (SIR) angiography that allows us to obtain a collection of several angiograms within the same acquisition time previously required to obtain a single image. In basic SIR, a single readout is performed after the tagging inversion pulse. In multiple-readout SIR, a succession of readout pulses is applied following the inversion pulse. By varying the gradients appropriately during the successive readouts, we can obtain a set of multiple projection-angle angiograms, or, by appropriately spacing the readouts throughout the cardiac cycle, we can obtain a set of time-resolved angiograms. This technique allows us to obtain additional spatial or temporal information without increasing total scan time. A sequence of increasing flip-angle read pulses is used to maintain a constant signal level across the images. A trade-off exists between SNR and the number of images acquired.

Blood Vessels↗

PubMed1993

Localized real-time velocity spectra determination.

The accurate measurement of flow velocity has long been a subject of NMR research. In the field of medical imaging, a variety of techniques primarily based on the principle of Fourier encoding have been described. Due to time constraints, necessary trade-offs exist between spatial versus velocity spectral resolution. In general, either the average velocity of individual pixels is displayed or velocity spectral determinations are made at the cost of spatial localization. The recent development of multidimensional excitation pulses makes spatial localization possible during the excitation phase of the pulse sequences. This approach, coupled with time varying gradient readout, can be used to obtain single-shot localized velocity spectra. Using these concepts, we have obtained in vivo real-time measurements of localized velocity spectra on our clinical imager.

Aorta↗

PubMed1995

Chemical shift imaging from simultaneous acquisition of a primary and a stimulated echo.

An imaging method is presented for obtaining chemical shift images from only one acquisition. Images are acquired with the same spatial resolution as in regular spin-echo imaging. The sequence is based on the simultaneous acquisition of a spin echo and a stimulated echo in a single pass. The use of 90 degrees radiofrequency pulse flip angles results in the same proton density, T2 and T1 weighting for the two echoes. Application of this sequence to chemical shift imaging is discussed for three fat suppression techniques (chopper, CHESS, and hybrid). Imaging was performed on phatoms and volunteers. The image quality was the same as those obtained by the chopper and the hybrid methods and the acquisition time was reduced by a factor of two.

Adipose Tissue↗

PubMed2006

Temporal diffusion spectroscopy: theory and implementation in restricted systems using oscillating gradients.

The theory of temporal diffusion spectra is reviewed. In contrast to q-space spectroscopy, which measures the displacement spectrum of spins in a spatial domain, the spectral density of the velocity correlation function (VCF) in the temporal domain is considered. It is demonstrated that casting diffusion in this domain may facilitate measurements of microscopic geometry and the decomposition of the diffusion signal into components due to disperse flow and restricted diffusion. An oscillating gradient (OG) method of diffusion spectroscopy was developed and implemented. Microscopic pore sizes, surface-to-volume ratios (S/Vs), and diffusion path tortuosities were extracted from model systems using this method. Cases are discussed in which this type of experiment may allow the characterization of pore geometry when spatial domain experiments fail. OGs may be combined with imaging sequences to map complex patterns of diffusion and flow. Moreover, scalar apparent diffusion coefficient (ADC) measurements in complex biological systems may be subtly dependent on specific pulse sequence parameters. Thus, scalar ADC measurements using gradient pulses with different frequency spectra may give different results. Conversely, the frequency dependence of motion-sensitizing gradient pulses may be exploited to deduce the origin of ADC changes.

Diffusion Magnetic Resonance Imaging↗

PubMed2006

Selective suppression of artifact-generating echoes in cine DENSE using through-plane dephasing.

In displacement-encoded imaging with stimulated echoes (DENSE), tissue displacement is encoded in the phase of the stimulated echo. However, three echoes generally contribute to the acquired signal (the stimulated echo, the complex conjugate of the stimulated echo, and an echo due to T(1) relaxation). It is usually desirable to suppress all except the stimulated echo, since otherwise the additional echoes will cause displacement measurement errors. Ideally, suppression of the artifact-generating echoes would be independent of time, T(1), and displacement-encoding frequency, and would not require additional acquisitions. In this study through-plane gradients were used to selectively dephase artifact-generating echoes without causing significant signal loss of the stimulated echo. A cine DENSE sequence was modified to include dephasing gradients and perform complementary spatial modulation of magnetization (CSPAMM). For single-acquisition cine DENSE using dephasing alone, artifact suppression was similar to CSPAMM with two acquisitions. The use of dephasing with CSPAMM required two acquisitions, but demonstrated greater artifact suppression than CSPAMM alone or dephasing alone.

Algorithms↗

PubMed1995

Characterization of the SpHE promoter that is spatially regulated along the animal-vegetal axis of the sea urchin embryo.

To understand how the maternally determined animal-vegetal polarity of the sea urchin embryo is established, we have begun to examine the regulatory apparatus of the gene encoding the Strongylocentrotus purpuratus hatching enzyme (SpHE). Previous studies have shown that the pattern of SpHE mRNA accumulation reflects the animal-vegetal developmental axis in that transcription is strongly upregulated during early cleavage in more animal blastomeres, but not in those around the maternally specified vegetal pole of the 16-cell embryo [Reynolds et al., Development 114, 769-786 (1992)]. Tests of SpHE promoter function in vivo using chloramphenicol acetyltransferase and beta-galactosidase enzymatic reporters define a regulatory region within several hundred nucleotides of the transcription initiation site. This region is sufficient to mediate both strong expression in the early blastula and spatially correct transcription. However, neither this region nor longer upstream sequences are sufficient to reproduce the transcriptional downregulation after very early blastula stage that is observed for endogenous genes. Biochemical assays of protein-DNA interactions within the regulatory region identify at least nine sites binding at least six different factors. These cis elements include Otx (an orthodenticle homologue), CCAAT, ets-related, and three unidentified motifs. Deletions and/or replacements of these cis-elements, alone and in combination, indicate that no single factor is essential for SpHE promoter activity, but instead that various combinations of subsets of these elements are capable of eliciting levels of transcription similar to those of the unaltered regulatory region. This density of regulatory elements is consistent with the intense transcription of endogenous SpHE genes during cleavage.

Animals↗

PubMed1996

Comparison of CH1 domains in different classes of murine antibodies.

The CH1 domains of antibodies belonging to the following five murine immunoglobulin (Ig) classes IgG1, IgG2a, IgG2b, IgG3 and IgA have been compared. The IgG CH1 domain structures are, as would be expected, similar overall, but show local conformational variations. When compared with IgG CH1 domain structures, the IgA CH1 domain displays several significant structural differences, which are a consequence of insertions/ deletions and specific structural constraints. In regions of structural differences in the IgG CH1 domains, the spatial correspondence of residues is not reflected by conventional (Kabat) sequence number. Thus the sequence alignment and numbering for CH1 domains has been revised to be consistent with the three-dimensional alignments.

Amino Acid Sequence↗

PubMedDate not supplied

Fold change in evolution of protein structures.

Typically, protein spatial structures are more conserved in evolution than amino acid sequences. However, the recent explosion of sequence and structure information accompanied by the development of powerful computational methods led to the accumulation of examples of homologous proteins with globally distinct structures. Significant sequence conservation, local structural resemblance, and functional similarity strongly indicate evolutionary relationships between these proteins despite pronounced structural differences at the fold level. Several mechanisms such as insertions/deletions/substitutions, circular permutations, and rearrangements in beta-sheet topologies account for the majority of detected structural irregularities. The existence of evolutionarily related proteins that possess different folds brings new challenges to the homology modeling techniques and the structure classification strategies and offers new opportunities for protein design in experimental studies.

Amino Acid Sequence↗

PubMed1998

Differential activation of right superior parietal cortex and intraparietal sulcus by spatial and nonspatial attention.

Neuropsychological and functional neuroimaging studies have implicated the right posterior parietal cortex (PPC) in human spatial attention. We tested the hypothesis that this area is also involved in nonspatial aspects of attention and working memory using positron emission tomography in healthy volunteers. In an initial experiment, digits were presented in pseudo-random spatial locations, and subjects attended either to locations or digits in order to detect single targets (attention condition) or to sequences of stimuli (working memory (WM) condition). Right superior parietal cortex (BA7) and intraparietal sulcus (IPS) were active during both spatial (locations) and nonspatial (digits) tasks compared to rest, although more so for the former. Additionally, right PPC was activated to an even greater extent during tests of WM than of attention, especially for tests of spatial WM. There were no differences in activation of dorsolateral prefrontal cortex in the spatial versus nonspatial versions of the task, contrary to many previous studies. A follow-up experiment which presented abstract objects in a fixed, central location confirmed that right IPS was active during tests of nonspatial attention and also that this activation is not due to incidental spatial representation of digit stimuli. However, BA7 was not activated by this nonspatial, nondigit attentional task. Overall, these data suggest first that right IPS is recruited for both nonspatial and spatial attention and WM. Second, right BA7 is recruited specifically for spatial (both direct and indirect) forms of attentional processing. Finally, PPC activations in spatial WM tasks are likely to be due to a combination of spatial perception, attention, and WM, rather than to any of these individually.

Adult↗

PubMed1993

A complex ensemble of cis-regulatory elements controls the expression of a Vicia faba non-storage seed protein gene.

We have identified cis-regulatory elements within the 5'-upstream region of a Vicia faba non-storage seed protein gene, called usp, by studying the expression of usp-promoter deletion fragments fused to reporter genes in transgenic tobacco seeds. 0.4 kb of usp upstream sequence contain at least six, but probably more, distinct cis-regulatory elements which are responsible for seemingly all quantitative, spatial and temporal aspects of expression. Expression-increasing and -decreasing elements are interspersed and include an AT-rich sequence, a G-box element and a CATGCATG motif. The latter acts as a negative element in contrast to what has been found for the same motif in legumin- and vicilin-type seed storage protein gene promoters. Seed specificity of expression is mainly determined by the -68/+51 region which confers, however, only very low levels of expression. The data support the combinatorial model of promoter function.

Base Sequence↗

PubMed1983

Ethanol and behavioral variability in the radial-arm maze.

Ethanol (0.75, 1.5, 2.0 g/kg ethyl alcohol) consistently and profoundly narrowed three independent dimensions of behavioral variability (BV) exhibited by rats in an eight-arm radial maze. Thie was true for all doses except the lowest. Rats were run with a replacement procedure wherein rewards (two food pellets) were replaced after they were taken. With no constraints against where, how, or by what route rewards could be taken, the three indices of spontaneous BV recorded were the number of different arms chosen, the sequence of visitation, and instances of deviations from goal-directed activity. The behavior of saline and low-dose groups was widely variable in form and place; and the sequence of behavior was relatively unpredictable from trial to trial and from session to session. Medium and high doses of ethanol exerted a marked organizing influence on behavior. Superfluous topographies were eliminated, sequences became highly, and in many cases perfectly predictable, and spatial BV declined. The considerable promotion of stereotypy by ethanol helps to explain many effects of the drug, e.g., how the drug can in some instances impair, and in others facilitate performance. We propose that the scores from tasks whose mastery entails repetition, few topographies, and rigid structure will be improved by ethanol, but that those requiring change and the sampling of new strategies will be impaired.

Animals↗