Proteomic identification of all plastid-specific ribosomal proteins in higher plant chloroplast 30S ribosomal subunit.

Six ribosomal proteins are specific to higher plant chloroplast ribosomes [Subramanian, A.R. (1993) Trends Biochem. Sci.18, 177-180]. Three of them have been fully characterized [Yamaguchi, K., von Knoblauch, K. & Subramanian, A. R. (2000) J. Biol. Chem. 275, 28455-28465; Yamaguchi, K. & Subramanian, A. R. (2000) J. Biol. Chem. 275, 28466-28482]. The remaining three plastid-specific ribosomal proteins (PSRPs), all on the small subunit, have now been characterized (2D PAGE, HPLC, N-terminal/internal peptide sequencing, electrospray ionization MS, cloning/ sequencing of precursor cDNAs). PSRP-3 exists in two forms (alpha/beta, N-terminus free and blocked by post-translational modification), whereas PSRP-2 and PSRP-4 appear, from MS data, to be unmodified. PSRP-2 contains two RNA-binding domains which occur in mRNA processing/stabilizing proteins (e.g. U1A snRNP, poly(A)-binding proteins), suggesting a possible role for it in the recruiting of stored chloroplast mRNAs for active protein synthesis. PSRP-3 is the higher plant orthologue of a hypothetical protein (ycf65 gene product), first reported in the chloroplast genome of a red alga. The ycf65 gene is absent from the chloroplast genomes of higher plants. Therefore, we suggest that Psrp-3/ycf65, encoding an evolutionarily conserved chloroplast ribosomal protein, represents an example of organelle-to-nucleus gene transfer in chloroplast evolution. PSRP-4 shows strong homology with Thx, a small basic ribosomal protein of Thermus thermophilus 30S subunit (with a specific structural role in the subunit crystallographic structure), but its orthologues are absent from Escherichia coli and the photosynthetic bacterium Synechocystis. We would therefore suggest that PSRP-4 is an example of gene capture (via horizontal gene transfer) during chloro-ribosome emergence. Orthologues of all six PSRPs are identifiable in the complete genome sequence of Arabidopsis thaliana and in the higher plant expressed sequence tag database. All six PSRPs are nucleus-encoded. The cytosolic precursors of PSRP-2, PSRP-3, and PSRP-4 have average targeting peptides (62, 58, and 54 residues long), and the mature proteins are of 196, 121, and 47 residues length (molar masses, 21.7, 13.8 and 5.2 kDa), respectively. Functions of the PSRPs as active participants in translational regulation, the key feature of chloroplast protein synthesis, are discussed and a model is proposed.     

Periodic breathing in the mouse.

The hypothesis was that unstable breathing might be triggered by a brief hypoxia challenge in C57BL/6J (B6) mice, which in contrast to A/J mice are known not to exhibit short-term potentiation; as a consequence, instability of ventilatory behavior could be inherited through genetic mechanisms. Recordings of ventilatory behavior by the plethsmography method were made when unanesthetized B6 or A/J animals were reoxygenated with 100% O(2) or air after exposure to 8% O(2) or 3% CO(2)-10% O(2) gas mixtures. Second, we examined the ventilatory behavior after termination of poikilocapnic hypoxia stimuli in recombinant inbred strains derived from B6 and A/J animals. Periodic breathing (PB) was defined as clustered breathing with either waxing and waning of ventilation or recurrent end-expiratory pauses (apnea) of > or = 2 average breath durations, each pattern being repeated with a cycle number > or = 3. With the abrupt return to room air from 8% O(2), 100% of the 10 B6 mice exhibited PB. Among them, five showed breathing oscillations with apnea, but none of the 10 A/J mice exhibited cyclic oscillations of breathing. When the animals were reoxygenated after 3% CO(2)-10% O(2) challenge, no PB was observed in A/J mice, whereas conditions still induced PB in B6 mice. (During 100% O(2) reoxygenation, all 10 B6 mice had PB with apnea.) Expression of PB occurred in some but not all recombinant mice and was not associated with the pattern of breathing at rest. We conclude that differences in expression of PB between these strains indicate that genetic influences strongly affect the stability of ventilation in the mouse.     

Suppression of atrial natriuretic peptide/natriuretic peptide receptor-A-mediated signaling upregulates angiotensin-II-induced collagen synthesis in adult cardiac fibroblasts

Cardiac hormone atrial natriuretic peptide (ANP) and its receptor natriuretic peptide receptor-A (NPR-A) system acts as an intrinsic negative regulator of abnormal extracellular matrix (ECM) remodeling in the heart. However, the underlying mechanism by which ANP/NPR-A system opposes the ECM remodeling in the diseased heart is not well understood. Here, we investigated the anti-fibrotic mechanism of ANP/NPR-A in fibrotic agonist Angiotensin- II (ANG II)-treated adult cardiac fibroblast (CF) cells. Normal and NPR-A-suppressed adult CF cells were treated with ANG II (10^?7 M) in the presence and absence of ANP (10^?8 M) for 24 h. Total collagen concentration, activity and expression of MMP-2 and MMP-9, and nuclear translocation of Nuclear factor-kappaB (NF-κB-p50) were studied. NPR-A-suppressed adult CF cells exhibited a more pronounced increase in collagen production, ROS generation, and NF-κB-p50 nuclear translocation as compared to adult CF cells treated with agonist alone. ANP co-treatment significantly reverses the agonist-induced above changes in normal adult CF cells, while it failed to reverse the agonist-induced collagen synthesis in the NPR-A-suppressed adult CF cells. The cGMP analog (8-bromo-cGMP) treatment significantly attenuated the agonist-induced collagen synthesis both in normal and NPR-A-suppressed adult cells. The results of this study suggest that ANP/NPR-A signaling system antagonizes the agonist-induced collagen synthesis via suppressing the activities of MMP-2, MMP-9, ROS generation, and NF-κB nuclear translocation mechanism.     

Endocrine response in rabbits immunized with native versus deglycosylated porcine zona pellucida antigens

Previous studies evaluating porcine zona pellucida antigens for immunocontraceptive purposes have in some cases revealed altered ovarian function in association with antibody response. This study was undertaken in an attempt to identify zona immunogens that do not cause adverse endocrine effects. To this end, we investigated the effects of highly purified preparations of native and deglycosylated pig zona pellucida antigens on ovarian function and immune response in the rabbit. Thirty female rabbits were immunized, 5 per group, with 100 micrograms each of either 1) SIZP, solubilized isolated zonae pellucidae; 2) ZP3, a purified porcine zona preparation containing the two principle glycoproteins, ZP3 alpha and ZP3 beta, endo-beta-galactosidase-digested ZP3 glycoproteins (approximately 30% deglycosylated) termed 3) ZP3 alpha/EBGD and 4) ZP3 beta/EBGD; and chemically deglycosylated ZP3 alpha and ZP3 beta (greater than or equal to 92% deglycosylated), termed 5) ZP3 alpha/DG and 6) ZP3 beta/DG. Rabbits injected with saline (n = 2) or Freund's adjuvant alone (n = 3) served as controls. Serum LH, FSH, estradiol, and progesterone were measured at 5-day intervals during seven 20-day cycles of hCG-induced pseudopregnancy over 42 wk. Anti-ZP3 titers, determined by RIA, developed in all treatment groups and correlated directly with carbohydrate content. Animals immunized with SIZP, ZP3, and ZP3 beta/EBGD showed a significant elevation of LH and FSH and a significant decline of peak progesterone levels by the fourth pseudopregnancy cycle. In contrast, animals immunized with ZP3 alpha/EBGD, ZP3 alpha/DG, and ZP3 beta/DG showed no significant elevations of gonadotropins and continued to display cyclic progesterone secretion in response to hCG.(ABSTRACT TRUNCATED AT 250 WORDS)     

The crystal at 5.5A resolution of an acid-protease from Rhizopus chinensis.

An electron density map for the pepsin-like enzyme from Rhizopus chinensis has been calculated at 5.5Å resolution. The molecular boundary has been defined and certain secondary structural details have been inferred. The molecule is bilobal and has a large cleft in which several inhibitors have been observed to bind.     

DIALIGN-TX: greedy and progressive approaches for segment-based multiple sequence alignment

Background  

DIALIGN-T is a reimplementation of the multiple-alignment program DIALIGN. Due to several algorithmic improvements, it produces significantly better alignments on locally and globally related sequence sets than previous versions of DIALIGN. However, like the original implementation of the program, DIALIGN-T uses a a straight-forward greedy approach to assemble multiple alignments from local pairwise sequence similarities. Such greedy approaches may be vulnerable to spurious random similarities and can therefore lead to suboptimal results. In this paper, we present DIALIGN-TX, a substantial improvement of DIALIGN-T that combines our previous greedy algorithm with a progressive alignment approach.