(Table 1) Gas tie points used to correlate NorthGRIP and Vostol records

North Atlantic sediment records (MD95-2042), Greenland (Greenland Ice Core Project (GRIP)) and Antarctica (Byrd and Vostok) ice core climate records have been synchronized over marine isotopic stage 3 (MIS 3) (64 to 24 kyr B.P.) (Shackleton et al., 2000). The resulting common timescale suggested that MD95-2042 d18Obenthic fluctuations were synchronous with temperature changes in Antarctica (dDice or d18Oice records). In order to assess the persistency of this result we have used here the recent Greenland NorthGRIP ice core covering the last glacial inception. We transfer the Antarctic Vostok GT4 timescale to NorthGRIP d18Oice and MD95-2042 d18Oplanktonic records and precisely quantify all the relative timing uncertainties. During the rapid warming of Dansgaard-Oeschger 24, MD95-2042 d18Obenthic decrease is in phase with d18Oplanktonic decrease and therefore with NorthGRIP temperature increase, but it takes place 1700 ± 1100 years after the Antarctic warming. Thus the present study reveals that the results obtained previously for MIS 3 cannot be generalized and demonstrates the need to improve common chronologies for marine and polar archives.

(Table 1) Gas tie points used to correlate the ice cores NGRIP and EDML

Water isotope records from the EPICA Dronning Maud Land (EDML) and the NorthGRIP ice cores have revealed a one to one coupling between Antarctic Isotope Maxima (AIM) and Greenland Dansgaard-Oeschger (DO) events back to 50 kyr. In order to explore if this north-south coupling is persistent over Marine Isotopic Stage 5 (MIS 5), a common timescale must first be constructed. Here, we present new records of d18O of O2 (d18Oatm) and methane (CH4) measured in the air trapped in ice from the EDML (68-147 kyr) and NorthGRIP (70-123 kyr) ice cores. We demonstrate that, through the period of interest, CH4 records alone are not sufficient to construct a common gas timescale between the two cores. Millennial-scale variations of d18Oatm are evidenced over MIS 5 both on the Antarctic and Greenland ice cores and are coupled to CH4 profiles to synchronise the NorthGRIP and EDML records. They are shown to be a precious tool for ice core synchronisation. With this new dating strategy, we produce the first continuous and accurate sequence of the north-south climatic dynamics on a common ice timescale for the last glacial inception and the first DO events of MIS 5, reducing relative dating uncertainties to an accuracy of a few centuries at the onset of DO events 24 to 20. This EDML-NorthGRIP synchronisation provides new firm evidence that the bipolar seesaw is a pervasive pattern from the beginning of the glacial period. The relationship between Antarctic warming amplitudes and their concurrent Greenland stadial duration highlights the particularity of DO event 21 and its Antarctic counterpart. Our results suggest a smaller Southern Ocean warming rate for this long DO event compared to DO events of MIS 3.

Synpolydactyly phenotypes correlate with size of expansions in HOXD13 polyalanine tract

Synpolydactyly (SPD) is a dominantly inherited congenital limb malformation. Typical cases have 3/4 finger and 4/5 toe syndactyly, with a duplicated digit in the syndactylous web, but incomplete penetrance and variable expressivity are common. The condition has recently been shown to be caused by expansions of an imperfect trinucleotide repeat sequence encoding a 15-residue polyalanine tract in HOXD13. We have studied 16 new and 4 previously published SPD families, with between 7 and 14 extra residues in the tract, to analyze the molecular basis for the observed variation in phenotype. Although there is no evidence of change in expansion size within families, even over six generations, there is a highly significant increase in the penetrance and severity of phenotype with increasing expansion size, affecting both hands (P = 0.012) and feet (P < 0.00005). Affected individuals from a family with a 14-alanine expansion, the largest so far reported, all have a strikingly similar and unusually severe limb phenotype, involving the first digits and distal carpals. Affected males from this family also have hypospadias, not previously described in SPD, but consistent with HOXD13 expression in the developing genital tubercle. The remarkable correlation between phenotype and expansion size suggests that expansion of the tract leads to a specific gain of function in the mutant HOXD13 protein, and has interesting implications for the role of polyalanine tracts in the control of transcription.

Intron positions correlate with module boundaries in ancient proteins

We analyze the three-dimensional structure of proteins by a computer program that finds regions of sequence that contain module boundaries, defining a module as a segment of polypeptide chain bounded in space by a specific given distance. The program defines a set of “linker regions” that have the property that if an intron were to be placed into each linker region, the protein would be dissected into a set of modules all less than the specified diameter. We test a set of 32 proteins, all of ancient origin, and a corresponding set of 570 intron positions, to ask if there is a statistically significant excess of intron positions within the linker regions. For 28-Å modules, a standard size used historically, we find such an excess, with P < 0.003. This correlation is neither due to a compositional or sequence bias in the linker regions nor to a surface bias in intron positions. Furthermore, a subset of 20 introns, which can be putatively identified as old, lies even more explicitly within the linker regions, with P < 0.0003. Thus, there is a strong correlation between intron positions and three-dimensional structural elements of ancient proteins as expected by the introns-early approach. We then study a range of module diameters and show that, as the diameter varies, significant peaks of correlation appear for module diameters centered at 21.7, 27.6, and 32.9 Å. These preferred module diameters roughly correspond to predicted exon sizes of 15, 22, and 30 residues. Thus, there are significant correlations between introns, modules, and a quantized pattern of the lengths of polypeptide chains, which is the prediction of the “Exon Theory of Genes.”

Unbinding forces of single antibody-antigen complexes correlate with their thermal dissociation rates

Point mutants of three unrelated antifluorescein antibodies were constructed to obtain nine different single-chain Fv fragments, whose on-rates, off-rates, and equilibrium binding affinities were determined in solution. Additionally, activation energies for unbinding were estimated from the temperature dependence of the off-rate in solution. Loading rate-dependent unbinding forces were determined for single molecules by atomic force microscopy, which extrapolated at zero force to a value close to the off-rate measured in solution, without any indication for multiple transition states. The measured unbinding forces of all nine mutants correlated well with the off-rate in solution, but not with the temperature dependence of the reaction, indicating that the same transition state must be crossed in spontaneous and forced unbinding and that the unbinding path under load cannot be too different from the one at zero force. The distance of the transition state from the ground state along the unbinding pathway is directly proportional to the barrier height, regardless of the details of the binding site, which most likely reflects the elasticity of the protein in the unbinding process. Atomic force microscopy thus can be a valuable tool for the characterization of solution properties of protein-ligand systems at the single molecule level, predicting relative off-rates, potentially of great value for combinatorial chemistry and biology.

β-Chemokines and neutralizing antibody titers correlate with sterilizing immunity generated in HIV-1 vaccinated macaques

One of the obstacles to AIDS vaccine development is the variability of HIV-1 within individuals and within infected populations, enabling viral escape from highly specific vaccine induced immune responses. An understanding of the different immune mechanisms capable of inhibiting HIV infection may be of benefit in the eventual design of vaccines effective against HIV-1 variants. To study this we first compared the immune responses induced in Rhesus monkeys by using two different immunization strategies based on the same vaccine strain of HIV-1. We then utilized a chimeric simian/HIV that expressed the envelope of a dual tropic HIV-1 escape variant isolated from a later time point from the same patient from which the vaccine strain was isolated. Upon challenge, one vaccine group was completely protected from infection, whereas all of the other vaccinees and controls became infected. Protected macaques developed highest titers of heterologous neutralizing antibodies, and consistently elevated HIV-1-specific T helper responses. Furthermore, only protected animals had markedly increased concentrations of RANTES, macrophage inflammatory proteins 1α and 1β produced by circulating CD8+ T cells. These results suggest that vaccine strategies that induce multiple effector mechanisms in concert with β-chemokines may be desired in the generation of protective immune responses by HIV-1 vaccines.

Trends in lipoplex physical properties dependent on cationic lipid structure, vehicle and complexation procedure do not correlate with biological activity

Using a group of structurally related cytofectins, the effects of different vehicle constituents and mixing techniques on the physical properties and biological activity of lipoplexes were systematically examined. Physical properties were examined using a combination of dye accessibility assays, centrifugation, gel electrophoresis and dynamic light scattering. Biological activity was examined using in vitro transfection. Lipoplexes were formulated using two injection vehicles commonly used for in vivo delivery (PBS pH 7.2 and 0.9% saline), and a sodium phosphate vehicle previously shown to enhance the biological activity of naked pDNA and lipoplex formulations. Phosphate was found to be unique in its effect on lipoplexes. Specifically, the accessible pDNA in lipoplexes formulated with cytofectins containing a γ-amine substitution in the headgroup was dependent on alkyl side chain length and sodium phosphate concentration, but the same effects were not observed when using cytofectins containing a β-OH headgroup substitution. The physicochemical features of the phosphate anion, which give rise to this effect in γ-amine cytofectins, were deduced using a series of phosphate analogs. The effects of the formulation vehicle on transfection were found to be cell type-dependent; however, of the formulation variables examined, the liposome/pDNA mixing method had the greatest effect on transgene expression in vitro. Thus, though predictive physical structure relationships involving the vehicle and cytofectin components of the lipoplex were uncovered, they did not extrapolate to trends in biological activity.

A neural correlate of syntactic encoding during speech production

Spoken language is one of the most compact and structured ways to convey information. The linguistic ability to structure individual words into larger sentence units permits speakers to express a nearly unlimited range of meanings. This ability is rooted in speakers' knowledge of syntax and in the corresponding process of syntactic encoding. Syntactic encoding is highly automatized, operates largely outside of conscious awareness, and overlaps closely in time with several other processes of language production. With the use of positron emission tomography we investigated the cortical activations during spoken language production that are related to the syntactic encoding process. In the paradigm of restrictive scene description, utterances varying in complexity of syntactic encoding were elicited. Results provided evidence that the left Rolandic operculum, caudally adjacent to Broca's area, is involved in both sentence-level and local (phrase-level) syntactic encoding during speaking.

Loss of oncogenic ras expression does not correlate with loss of tumorigenicity in human cells.

ras oncogenes are mutated in at variety of human tumors, which suggests that they play an important role in human carcinogenesis. To determine whether continued oncogenic ras expression is necessary to maintain the malignant phenotype, we studied the human fibrosarcoma cell line, HT1080, which contains one mutated and one wild-type N-ras allele. We isolated a variant of this cell line that no longer contained the mutated copy of the N-ras gene. Loss of mutant N-ras resulted in cells that displayed a less transformed phenotype characterized by a flat morphology, decreased growth rate, organized actin stress fibers, and loss of anchorage-independent growth. The transformed phenotype was restored following reintroduction of mutant N-ras. Although loss of the oncogenic N-ras drastically affected in vitro growth parameters, the variant remained tumorigenic in nude mice indicating that mutated N-ras expression is not necessary for maintenance of the tumorigenic phenotype. We confirmed this latter observation in colon carcinoma cell lines that have lost activated K-ras expression via targeted knockout of the mutant K-ras gene.