A longitudinal study of thyroid autoantibodies in pregnancy: the importance of test timing.

OBJECTIVE: Thyroid peroxidase antibodies (TPOAb) and thyroglobulin antibodies (TGAb) are frequently measured to investigate thyroid dysfunction in pregnancy. Despite the recognized fall of these autoantibodies in pregnancy, there is limited guidance on the timing of such testing. We assessed optimal test timing of TPOAb/TGAb for the detection of Hashimoto's thyroiditis and post-partum thyroid dysfunction (PPTD). DESIGN: Prospective longitudinal study with recruitment in Trimester 1. PATIENTS: Healthy women ≤13 weeks' gestation from Mercy Hospital for Women, a tertiary obstetric hospital in Melbourne. MEASUREMENTS: Serum TPOAb, TGAb, TSH and fT4 were measured at Trimester 1 (T1), Trimester 2(T2), Trimester 3(T3) and postpartum (PP) in each participant. Post-partum thyroid dysfunction (PPTD) was defined if TSH deviated from the assay's nonpregnant reference interval. Longitudinal random-effect logistic regression was used to investigate the association between time and positive/negative thyroid autoantibody status. RESULTS: Samples from 140 women at T1 (12·0: 10·3-13·0) (median: IQR weeks' gestation); 95 at T2 (24·3: 23·0-25·9), 79 at T3 (35·9: 34·8-36·7) and 83 at PP (12·4: 10·8-14·6 weeks post-partum) were attained. At T1, 13 (9%) and 15 (11%) women had positive TPOAb and TGAb, respectively. The odds of having a positive TPOAb were 96% lower at T2 [OR = 0·04 (95% CI: 0·02-0·8; P = 0·03)] and 97% lower at T3 [OR = 0·03 (95% CI: 0·001-0·6; P = 0·02)] than at T1. Similarly, the odds of having a positive TGAb were 99·4% lower [OR = 0·006 (95% CI: 0-0·3; P = 0·01)] at T2, and 99·5% lower [OR = 0·005 (95% CI: 0-0·4; P = 0·02)] at T3 than at T1. The ROC analysis diagnostic ORs for a positive TPOAb and/or TGAb to predict PPTD were 7·8 (95% CI: 2·2-27·6), 1·2 (95% CI: 0-8·9), 2·0 (95% CI: 0-16·8), and 12·2 (95% CI: 3·3-44·9) at T1, T2, T3 and post-partum, respectively. CONCLUSIONS: A significant proportion of pregnant women lose their thyroid autoantibody positivity after T1. The gestation-dependent loss of TPOAb/TGAb positivity and reduction in diagnostic accuracy for predicting PPTD limits the value of testing at T2 and T3.

Plant mitogen-activated protein kinase signaling cascades.

Mitogen-activated protein kinase (MAPK) cascades have emerged as a universal signal transduction mechanism that connects diverse receptors/sensors to cellular and nuclear responses in eukaryotes. Recent studies in plants indicate that MAPK cascades are vital to fundamental physiological functions involved in hormonal responses, cell cycle regulation, abiotic stress signaling, and defense mechanisms. New findings have revealed the complexity and redundancy of the signaling components, the antagonistic nature of distinct pathways, and the use of both positive and negative regulatory mechanisms.

NTDB: Thermodynamic Database for Nucleic Acids.

A new thermodynamic database for normal and modified nucleic acids has been developed. This Thermodynamic Database for Nucleic Acids (NTDB) includes sequence, structure and thermodynamic information as well as experimental methods and conditions. In this release, there are 1851 sequences containing both normal and modified nucleic acids. A user-friendly web-based interface has been developed to allow data searching under different conditions. Useful thermodynamic tools for the study of nucleic acids have been collected and linked for easy usage. NTDB is available at http://ntdb.chem.cuhk.edu.hk.

Complete nucleotide sequence of the Oenothera elata plastid chromosome, representing plastome I of the five distinguishable euoenothera plastomes.

We describe the 159,443-bp [corrected] sequence of the plastid chromosome of Oenothera elata (evening primrose). The Oe. elata plastid chromosome represents type I of the five genetically distinguishable basic plastomes found in the subsection Euoenothera. The genus Oenothera provides an ideal system in which to address fundamental questions regarding the functional integration of the compartmentalised genetic system characteristic of the eukaryotic cell. Its highly developed taxonomy and genetics, together with a favourable combination of features in its genetic structure (interspecific fertility, stable heterozygous progeny, biparental transmission of organelles, and the phenomenon of complex heterozygosity), allow facile exchanges of nuclei, plastids and mitochondria, as well as individual chromosome pairs, between species. The resulting hybrids or cybrids are usually viable and fertile, but can display various forms of developmental disturbance.

Functional analysis of oxidative stress-activated mitogen-activated protein kinase cascade in plants.

Despite the recognition of H(2)O(2) as a central signaling molecule in stress and wounding responses, pathogen defense, and regulation of cell cycle and cell death, little is known about how the H(2)O(2) signal is perceived and transduced in plant cells. We report here that H(2)O(2) is a potent activator of mitogen-activated protein kinases (MAPKs) in Arabidopsis leaf cells. Using epitope tagging and a protoplast transient expression assay, we show that H(2)O(2) can activate a specific Arabidopsis mitogen-activated protein kinase kinase kinase, ANP1, which initiates a phosphorylation cascade involving two stress MAPKs, AtMPK3 and AtMPK6. Constitutively active ANP1 mimics the H(2)O(2) effect and initiates the MAPK cascade that induces specific stress-responsive genes, but it blocks the action of auxin, a plant mitogen and growth hormone. The latter observation provides a molecular link between oxidative stress and auxin signal transduction. Finally, we show that transgenic tobacco plants that express a constitutively active tobacco ANP1 orthologue, NPK1, display enhanced tolerance to multiple environmental stress conditions without activating previously described drought, cold, and abscisic acid signaling pathways. Thus, manipulation of key regulators of an oxidative stress signaling pathway, such as ANP1/NPK1, provides a strategy for engineering multiple stress tolerance that may greatly benefit agriculture.

Suppression of auxin signal transduction by a MAPK cascade in higher plants.

The plant hormone auxin activates many early response genes that are thought to be responsible for diverse aspects of plant growth and development. It has been proposed that auxin signal transduction is mediated by a conserved signalling cascade consisting of three protein kinases: the mitogen-activated protein kinase (MAPK), MAPK kinase (MAPKK) and MAPKK kinase (MAPKKK). Here we show that a specific plant MAPKKK, NPK1, activates a MAPK cascade that leads to the suppression of early auxin response gene transcription. A mutation in the kinase domain abolishes NPK1 activity, and the presence of the carboxy-terminal domain diminishes the kinase activity. Moreover, the effects of NPK1 on the activation of a MAPK and the repression of early auxin response gene transcription are specifically eliminated by a MAPK phosphatase. Transgenic tobacco plants overexpressing the NPK1 kinase domain produced seeds defective in embryo and endosperm development. These results suggest that auxin sensitivity may be balanced by antagonistic signalling pathways that use a distinct MAPK cascade in higher plants.

Premature polyadenylation at multiple sites within a Bacillus thuringiensis toxin gene-coding region.

Some foreign genes introduced into plants are poorly expressed, even when transcription is controlled by a strong promoter. Perhaps the best examples of this problem are the cry genes of Bacillus thuringiensis (B.t.), which encode the insecticidal proteins commonly referred to as B.t. toxins. As a step toward overcoming such problems most effectively, we sought to elucidate the mechanisms limiting the expression of a typical B.t.-toxin gene, cryIA(c), which accumulates very little mRNA in tobacco (Nicotiana tabacum) cells. Most cell lines transformed with the cryIA(c) B.t.-toxin gene accumulate short, polyadenylated transcripts. The abundance of these transcripts can be increased by treating the cells with cycloheximide, a translation inhibitor that can stabilize many unstable transcripts. Using a series of hybridizations, reverse-transcriptase polymerase chain reactions, and RNase-H-digestion experiments, poly(A+) addition sites were identified in the B.t.-toxin-coding region corresponding to the short transcripts. A fourth polyadenylation site was identified using a chimeric gene. These results demonstrate for the first time to our knowledge that premature polyadenylation can limit the expression of a foreign gene in plants. Moreover, this work emphasizes that further study of the fundamental principles governing polyadenylation in plants will have basic as well as applied significance.

Characterization of primary lesions caused by the plastome mutator of Oenothera.

Oenothera plants homozygous for a recessive allele at the plastome mutator (pm) locus show non-Mendelian mutation frequencies that are 1000-fold higher than spontaneous levels. Characterization of RFLP sites in a collection of mutants indicates that insertion-deletion hot spots in the pm lines are defined by tandem direct repeats, implicating replication slippage or misalignment during recombination. Several sites known to contain very short direct repeats were examined, and all were found to have been targeted in one or more plants of the mutant collection. To determine if replication slippage was occurring, two oligo-A stretches in non-coding DNA were examined, and 3 of 12 plants were found to have an additional adenine in a 13-base track. To search for other mutations that would not be visible as restriction fragment length polymorphisms, PCR-amplification products of the psbB gene were digested with a restriction endonuclease, denatured, and examined for single-strand conformational polymorphisms. Among 21 mutants, one 4-bp insertion and one point mutation were identified in psbB. The discovery that the plastome mutator can cause base substitutions as well as repeat-mediated insertions and deletions points to a likely defect in a component of the cpDNA replication machinery.

Proliferation of direct repeats near the Oenothera chloroplast DNA origin of replication.

The spacer between the 16S and 23S rRNA genes of the chloroplast DNA has been implicated as an origin of replication in several species of plants. In the evening primrose, Oenothera, this site was found to vary greatly in size, with plastid genomes (plastomes) being readily distinguished. To determine whether plastome "strength" in transmission could be correlated with variation at oriB, the 16S rRNA-trnI spacer was sequenced from five plastomes. The size variation was found to be due to differential amplification (and deletion) of combinations of sequences belonging to seven families of direct repeats. From these comparisons, one short series of direct repeats and one region capable of forming a hairpin structure were identified as candidates for the factor that could be responsible for the differences between strong and weak plastome types. Ample sequence variation allowed phylogenetic inferences to be made about the relationships among the plastomes. Phylogenetic trees also could be constructed for most of the families of direct repeats. The amplifications and deletions of repeats that account for the size variation at oriB are proposed to have occurred through extensive replication slippage at this site.

Extramedullary plasmacytoma of the head and neck.

Extramedullary plasmacytoma (EMP) is an uncommon malignant plasma cell neoplasm which usually presents as soft tissue mass in the head and neck. We retrospectively reviewed 7 cases with EMP of the head and neck between 1978 and 1992. All patients were male. Their ages ranged from 27 to 66 years. M-protein was identified in 5 patients (IgG-kappa in 3, IgG-lambda in 1 and Lambda light chain in 1). All but one were treated with local irradiation. Three patients received chemotherapy. All patients have been alive for 37+ to 116+ months since the diagnosis of EMP. One patient had a cervical node recurrence 9 years after local irradiation and the other one who refused local irradiation but received chemotherapy developed multiple myeloma 5 years later. Based on our experience and the review of the literature, it is recommended that irradiation to the primary sites and the involved cervical nodes is the treatment of choice. Additional chemotherapy may be considered in patients with disease progression, recurrence, or dissemination. The M-protein is a useful parameter to assess tumor control or disease progression.

Plastome-genome interactions affect plastid transmission in Oenothera.

Plastids of Oenothera, the evening primrose, can be transmitted to the progeny from both parents. In a constant nuclear background, the frequency of biparental plastid transmission is determined by the types of plastid genomes (plastomes) involved in the crosses. In this study, the impact of nuclear genomes on plastid inheritance was analyzed. In general, the transmission efficiency of each plastome correlated strongly with its compatibility with the nuclear genome of the progeny, suggesting that plastome-genome interactions can influence plastid transmission by affecting the efficiency of plastid multiplication after fertilization. Lower frequencies of plastid transmission from the paternal side were observed when the pollen had poor vigor due to an incompatible plastome-genome combination, indicating that plastome-genome interactions may also affect the input of plastids at fertilization. Parental traits that affect the process of fertilization can also have an impact on plastid transmission. Crosses using maternal parents with long styles or pollen with relatively low growth capacity resulted in reduced frequencies of paternal plastid transmission. These observations suggest that degeneration of pollen plastids may occur as the time interval between pollination and fertilization is lengthened.

Electron microscopic localization of replication origins in Oenothera chloroplast DNA.

The origins of chloroplast DNA (cpDNA) replication were mapped in two plastome types of Oenothera in order to determine whether variation in the origin of cpDNA replication could account for the different transmission abilities associated with these plastomes. Two pairs of displacement loop (D-loop) initiation sites were observed on closed circular cpDNA molecules by electron microscopy. Each pair of D-loops was mapped to the inverted repeats of the Oenothera cpDNA by the analysis of restriction fragments. The starting points of the two adjacent D-loops are approximately 4 kb apart, bracketing the 16S rRNA gene. Although there are small DNA length variations near one of the D-loop initiation sites, no apparent differences in the number and the location of replication origins were observed between plastomes with the highest (type I) and lowest (type IV) transmission efficiencies.

Recombination between chloroplast DNAs does not occur in sexual crosses of Oenothera.

Crosses of Oenothera result in the transmission of chloroplasts from both parents to their offspring. In spite of this biparental inheritance, no wild-type recombinants were recovered from crosses between different chloroplast mutants. Since more than 7500 progeny were examined, the results indicate that recombination between the chloroplast DNAs of higher plants must be a very rare event.