Paul J. Hagerman

Bio: Paul J. Hagerman is an academic researcher from University of California, Davis. The author has contributed to research in topics: FMR1 & Fragile X syndrome. The author has an hindex of 93, co-authored 281 publications receiving 26411 citations. Previous affiliations of Paul J. Hagerman include University of Colorado Boulder & Boston Children's Hospital.

Flexibility of DNA

Abstract: Both microscopic and macroscopic models of DNA flexibility should lead to the same quantitative description of the elastic properties of the DNA helix. This belief is reinforced by the fact that essentially all experimental (solution) studies to date support the macroscopic, elastic model. The performance of microscopic models can therefore be checked by their ability to produce the correct macroscopic quantities (P and C). To most carefully address the influence of such factors as base sequence, DNA damage, and drug or protein interaction on the flexibility of DNA, methods are required that are most sensitive for DNA molecules of less than 500-1000 bp. The use of molecules in this size range will maximize the signal due to the structural alteration as well as facilitate the construction of DNA sequences of any desired arrangement. I have emphasized three such methods and summarized their strengths and weaknesses; however, their concurrent application to the determination of DNA flexibility provides an important check of self-consistency. These studies have indicated that the persistence length of DNA in buffers of moderate salt concentration is 450-500 A. Synthetic DNA is now readily available, and many procedures for the construction and cloning of DNA molecules of defined length and sequence (107-108a) are in common use. The availability of restriction fragments of precisely defined length has transformed the study of the physical (particularly hydrodynamic) properties of such molecules, since the hitherto pervasive problem of length polydispersity has been eliminated. Sheared, sonicated, or otherwise abused calf thymus (or other) DNAs should no longer be considered acceptable materials for physical studies. Many studies of bending and torsional fluctuations in DNA have been excluded from this discussion because the DNA samples used were not precisely defined. The torsional elastic constant of DNA has been fairly well established as approximately 3.0 x 10(-19) erg-cm, mainly through a combination of elegant theoretical and experimental studies of topoisomer distributions in circular DNA molecules. The other general approach to the determination of the torsional elastic constant, luminescence decay, is still burdened by the poor characterization of the DNA used in many of the experimental studies as well as by some continued theoretical uncertainties.(ABSTRACT TRUNCATED AT 400 WORDS)

Intention tremor, parkinsonism, and generalized brain atrophy in male carriers of fragile X.

Abstract: The authors report five elderly men with the fragile X premutation who had a progressive action tremor associated with executive function deficits and generalized brain atrophy. These individuals had elevated fragile X mental retardation 1 gene (FMR1) messenger RNA and normal or borderline levels of FMR1 protein. The authors propose that elevations of FMR1 messenger RNA may be causative for a neurodegenerative syndrome in a subgroup of elderly men with the FMR1 premutation.

Elevated levels of FMR1 mRNA in carrier males : A new mechanism of involvement in the fragile-X syndrome

Abstract: Summary Fragile-X syndrome is a trinucleotide-repeat–expansion disorder in which the clinical phenotype is believed to result from transcriptional silencing of the fragile-X mental retardation 1 ( FMR1 ) gene as the number of CGG repeats exceeds ∼200. For premutation alleles (∼55–200 repeats), no abnormalities in FMR1 -gene expression have been described, despite growing evidence of clinical involvement in premutation carriers. To address this (apparent) paradox, we have determined, for 16 carrier males (55–192 repeats), the relative levels of leukocyte FMR1 mRNA, by use of automated fluorescence-detection reverse transcriptase–PCR, and the percent of lymphocytes that are immunoreactive for FMR1 protein (FMRP). For some alleles with >100 repeats, there was a reduction in the number of FMRP-positive cells. Unexpectedly, FMR1 mRNA levels were elevated at least fivefold within this same range. No significant increase in FMR1 mRNA stability was observed in a lymphoblastoid cell line (160 repeats) derived from one of the carrier males, suggesting that the increased message levels are due to an increased rate of transcription. Current results support a mechanism of involvement in premutation carriers, in which reduced translational efficiency is at least partially compensated through increased transcriptional activity. Thus, diminished translational efficiency may be important throughout much of the premutation range, with a mechanistic switch occurring in the full-mutation range as the FMR1 gene is silenced.

Fragile X Premutation Tremor/Ataxia Syndrome: Molecular, Clinical, and Neuroimaging Correlates

Abstract: We present a series of 26 patients, all >50 years of age, who are carriers of the fragile X premutation and are affected by a multisystem, progressive neurological disorder. The two main clinical features of this new syndrome are cerebellar ataxia and/or intention tremor, which were chosen as clinical inclusion criteria for this series. Other documented symptoms were short-term memory loss, executive function deficits, cognitive decline, parkinsonism, peripheral neuropathy, lower limb proximal muscle weakness, and autonomic dysfunction. Symmetrical regions of increased T2 signal intensity in the middle cerebellar peduncles and adjacent cerebellar white matter are thought to be highly sensitive for this neurologic condition, and their presence is the radiological inclusion criterion for this series. Molecular findings include elevated mRNA and low-normal or mildly decreased levels of fragile X mental retardation 1 protein. The clinical presentation of these patients, coupled with a specific lesion visible on magnetic resonance imaging and with neuropathological findings, affords a more complete delineation of this fragile X premutation–associated tremor/ataxia syndrome and distinguishes it from other movement disorders.

Penetrance of the Fragile X–Associated Tremor/Ataxia Syndrome in a Premutation Carrier Population

Abstract: ContextPremutation expansions (55-200 CGG repeats) of the fragile X mental retardation 1 (FMR1) gene are frequent in the general population, with estimated prevalences of 1 per 259 females and 1 per 813 males. Several articles have recently described the presence of late-onset neurological symptoms in male carriers of premutation (FMR1) alleles. The main clinical features described in this newly identified syndrome are cerebellar ataxia and intention tremor. Additional documented symptoms include short-term memory loss, executive functional deficits, cognitive decline, parkinsonism, peripheral neuropathy, lower-limb proximal muscle weakness, and autonomic dysfunction.ObjectiveTo study the penetrance of the fragile X–associated tremor/ataxia syndrome (FXTAS) among premutation carriers.Design, Setting, and ParticipantsFamily-based study of 192 individuals (premutation carriers and controls) whose families belong to the Northern or Southern California Fragile X Associations. Data were collected (March 2002-April 2003) through a survey and a standardized neurological examination, which was videotaped and subsequently scored in a blinded fashion.Main Outcome MeasuresPenetrance of intention tremor and ataxia among adult carriers (aged ≥50 years) of premutation expansions of the FMR1 gene.ResultsData from the survey of 192 individuals demonstrated an age-related penetrance of the combination of reported intention tremor and gait ataxia in male carriers (17%, 38%, 47%, and 75% [lower-bound estimates] for participants aged 50-59, 60-69, 70-79, and ≥80 years, respectively). The male carrier group had an age-adjusted 13-fold increased risk (95% confidence interval, 3.9-25.4; P = .003) of combined intention tremor and gait ataxia when compared with male controls. The clinical examination data from 93 individuals demonstrated that male carriers experienced more difficulties on each of 3 standardized neurological rating scales compared with controls (P<.05). Female carrier scores were also higher than those of female controls (P<.05) on 2 of the 3 neurological rating scales, but no participant was identified with probable or definite FXTAS.ConclusionsThe study demonstrates that older male carriers of premutation alleles of the FMR1 gene are at high risk of developing FXTAS. Since male premutation carriers are relatively common in the general population, older men with ataxia and intention tremor should be screened for the FMR1 mutation, especially if these signs are accompanied by parkinsonism, autonomic dysfunction, or cognitive decline, regardless of family history.

Molecular Beacons: Probes that Fluoresce upon Hybridization

Abstract: We have developed novel nucleic acid probes that recognize and report the presence of specific nucleic acids in homogeneous solutions. These probes undergo a spontaneous fluorogenic conformational change when they hybridize to their targets. Only perfectly complementary targets elicit this response, as hybridization does not occur when the target contains a mismatched nucleotide or a deletion. The probes are particularly suited for monitoring the synthesis of specific nucleic acids in real time. When used in nucleic acid amplification assays, gene detection is homogeneous and sensitive, and can be carried out in a sealed tube. When introduced into living cells, these probes should enable the origin, movement, and fate of specific mRNAs to be traced.

Nonhybrid, finished microbial genome assemblies from long-read SMRT sequencing data

Abstract: We present a hierarchical genome-assembly process (HGAP) for high-quality de novo microbial genome assemblies using only a single, long-insert shotgun DNA library in conjunction with Single Molecule, Real-Time (SMRT) DNA sequencing. Our method uses the longest reads as seeds to recruit all other reads for construction of highly accurate preassembled reads through a directed acyclic graph-based consensus procedure, which we follow with assembly using off-the-shelf long-read assemblers. In contrast to hybrid approaches, HGAP does not require highly accurate raw reads for error correction. We demonstrate efficient genome assembly for several microorganisms using as few as three SMRT Cell zero-mode waveguide arrays of sequencing and for BACs using just one SMRT Cell. Long repeat regions can be successfully resolved with this workflow. We also describe a consensus algorithm that incorporates SMRT sequencing primary quality values to produce de novo genome sequence exceeding 99.999% accuracy.

Real-Time DNA Sequencing from Single Polymerase Molecules

Abstract: We present single-molecule, real-time sequencing data obtained from a DNA polymerase performing uninterrupted template-directed synthesis using four distinguishable fluorescently labeled deoxyribonucleoside triphosphates (dNTPs). We detected the temporal order of their enzymatic incorporation into a growing DNA strand with zero-mode waveguide nanostructure arrays, which provide optical observation volume confinement and enable parallel, simultaneous detection of thousands of single-molecule sequencing reactions. Conjugation of fluorophores to the terminal phosphate moiety of the dNTPs allows continuous observation of DNA synthesis over thousands of bases without steric hindrance. The data report directly on polymerase dynamics, revealing distinct polymerization states and pause sites corresponding to DNA secondary structure. Sequence data were aligned with the known reference sequence to assay biophysical parameters of polymerization for each template position. Consensus sequences were generated from the single-molecule reads at 15-fold coverage, showing a median accuracy of 99.3%, with no systematic error beyond fluorophore-dependent error rates.