Distribution and evolution of C4 syndrome inRhynchospora (Rhynchosporeae-Cyperaceae)

In order to elucidate the evolution of C₄ syndrome, the taxonomic relationships, leaf anatomy, and ecological and global distribution of C₃ and C₄ species in the genusRhynchospora were investigated. The anatomical observation for 181 species revealed that 26 C₄ species occurred within theCapitatae group of the subgenusHaplostyleae, a natural group showing highly advanced morphological characteristics, together with several C₃ species. In spite of there being rather few C₄ species, they possessed two kinds of Kranz anatomical structure differing from each other in the location of Kranz cells. Some C₃ species ofCapitatae showed radial arrangement in mesophyll cells surrounding vascular bundles, which is distinguished from typical non-Kranz anatomy. The C₄ species extended their ecological ranges from wet habitats to dry savanna grasslands, while the C₃ species showed the best development in wet habitats. The C₃ species were widespread from tropical to temperate regions with partial range extension into subarctic regions of both hemispheres, showing conspicuously high concentration of species in the New World, but being absent from arid climatic regions. The C₄ species were distributed mostly in tropics and subtropics, showing two separate distributional centers in South and Central America and in Tropical Asia and Australia. The range of C₄ species was nearly completely included in the C₃ range. In conclusion, it seems that inRhynchospora the C₄ syndrome evolved relatively recently, and arose in at least two separate phylogenetic trends in the tropics and the subtropics, more probably in the Neotropics.     

Inhibition of tyrosine autophosphorylation of the solubilized insulin receptor by an insulin-stimulating peptide derived from bovine serum albumin

A polypeptide from a tryptic digest of bovine serum albumin potentiates glucose oxidation stimulated by insulin in isolated rat adipocytes. We studied whether this effect is related to a modification of the insulin receptor kinase. In a solubilized rat adipocytes receptor system, the peptide caused dose-dependent inhibition of the stimulation by insulin of phosphorylation of the 95,000 dalton subunit of insulin receptor. The peptide also inhibited stimulation by vanadate of tyrosine autophosphorylation of the beta subunit of the receptor, though it enhanced vanadate-stimulated glucose oxidation. During the phosphorylation reaction, no phosphorylated forms of the peptide could be detected. The peptide had no effect on dephosphorylation of the phosphorylated beta subunit of the insulin receptor. These results strongly suggest that the inhibition of phosphorylation by the peptide is due not to either simple substrate competition or activation of phosphoprotein phosphatase, but to specific inhibition of tyrosine-specific protein kinase.     

Properties of carboxymethylated cross-linked hemoglobin A

The selective carboxymethylation of the N-terminal amino groups of hemoglobin A with glyoxylic acid and sodium cyanoborohydride has been studied as a function of the state of ligation of hemoglobin. The N-terminal residues have been established as the primary sites of reaction by peptide mapping of the tryptic digest of each chain and subsequent amino acid analysis of the modified peptides. With oxyhemoglobin, the desired derivatives with a carboxymethyl group at the N-terminal of either or both chains amounted to 55% [Di Donato, A., Fantl, W. J., Acharya, A. S., & Manning, J. M. (1983) J. Biol. Chem. 258, 11890-11895]. In the present study it is shown that with deoxyhemoglobin the amount of the desired derivative is increased to 75%. The oxygen equilibrium curve of hemoglobin A carboxymethylated on its four N-terminal residues [0.5 mM as tetramer in 50 mM [bis(2-hydroxyethyl)amino]tris(hydroxymethyl)methane (Bis-Tris), pH 7.5, 37 degrees C] had a P50 value of 30 mmHg (Hill coefficient n = 2.8, alkaline Bohr value = 0.4) compared to a P50 of 9 mmHg for unmodified hemoglobin under the same conditions (n = 2.5, alkaline Bohr value = 0.5). In carboxymethylated oxyhemoglobin A, cross-linked with the mild agent glycolaldehyde for 3.5 h, there was 85% of Mr 64,000 species and 15% of Mr 128,000 or higher species. For the former, the extent of cross-linking between two subunits was 19%. For the latter, there was 29% of two cross-linked subunits and 13% of three cross-linked subunits. Termination of cross-linking, which may be desirable in some circumstances, can be successfully achieved with isonicotinic acid hydrazide.(ABSTRACT TRUNCATED AT 250 WORDS)     

Conformation between the substrate-binding site and heme of cytochrome P-450 studied by excitation energy transfer

The conformation between the substrate-binding site and heme of cytochrome P-450 was studied by excitation energy transfer. Cytochrome P-450 was obtained from the hepatic microsomes of polychlorinated biphenyl-treated male rats, and ten polycyclic aromatic hydrocarbons were used as substrates. The energy transfer from the substrate to the heme of the enzyme was measured according to the F?rster equation. On the basis of the assumption that the substrates are bound at different positions in the plane of the same substrate-binding site, the position of the heme in relation to the substrate-binding site was determined in solution and in the presence of synthetic phospholipid. The results demonstrated that the distance between the substrate-binding site and the heme of cytochrome P-450 was greater when the enzyme was incorporated into micelles of phospholipid than when in solution, and that the conformational relationship of the substrate-binding site towards the heme was changed by an angle of 21 degrees on incorporation of the enzyme into phospholipid micelles.     

Extensin Secreted into the Culture Medium by Tobacco Cells I. Purification and Some Properties

The medium of 12-day-old culturs of tobacco cells (Nicotianatabacum L., var Xanthi; line XD-6S) contain c.a. 160mg/literof protein, of which 14% of the constituent ami no acids werefound to be hydroxyproline. By sequential column chromatographiesand CsCl density-gradient centrifugation, a basic hydroxyproline-richglycoprotein was purified from the medium and found to havean amino acid composition typical of extensin; with a high levelof hydroxyproline (33mole%), tyrosine (13%), and lysine (14%).The glycoprotein contained 42% (w/w) of sugars, among whicharabinose was the major component (85%). The proportion of thisextensin in the proteins in the culture medium was estimatedto be much higher than that of arabino-galactan protein (about5 times higher) on a protein basis, with extensin comprisingbetween 25% and 41%, and probably about 37% of the proteinsin the medium. (Received September 19, 1988; Accepted December 26, 1988)     

Human thymocytes do not respond to interleukin-2 after removal of mature "bright" CD5 positive cells

Interleukin-2 receptors (IL-2R) are expressed on minor populations of immature and mature human thymocytes. These studies were designed to determine if immature T cells could respond to the mitogen phytohemagglutinin (PHA-P) plus IL-2 in vitro by increasing the expression of IL-2R and by proliferation. Using monoclonal antibodies to CD5 and magnetic immunobeads we were able to remove all mature, "bright" CD5+ cells from nylon wool-purified thymocytes and to obtain less mature cells which consisted almost completely of cells with the CD4+CD8+ phenotype. These immature cells were mostly "dim" CD5+ and less than 5% CD5- and a small percentage expressed the IL-2R. After culture in serum-free medium with PHA-P, these cells showed only a slight increase in the percentage of IL-2R+ cells and the addition of IL-2 did not increase the percentage of IL-2R+ cells and no proliferation was observed. Unseparated, nylon wool-purified thymocytes contained 14% bright CD5+ cells. These bright CD5+ cells had a mature phenotype of CD4+CD8- (52%) and CD4-CD8+ (27%) cells. A small percentage of these cells were IL-2R+. These bright CD5+IL-2R+ cells were predominantly mature CD4+CD8- cells as measured by three-color flow cytometry. After culture with PHA-P and IL-2, the percentage of IL-2R+ cells increased and they were now found not only on CD4+CD8- but also on CD4-CD8+ and on CD4+CD8+ cells. IL-2 plus PHA-P increased proliferation of these cells as compared to those cultured in medium with PHA-P without IL-2. Thus, we show that human immature thymocytes in contrast to mature thymocytes are not responsive to IL-2 as measured by a lack of IL-2R expression and proliferation. These data indicate that mature thymocytes can express a functional high affinity receptor for IL-2 and suggest that immature thymocytes may not possess a (functional) p75 chain of the IL-2R.     

Effects of sodium depletion and orthostasis on plasma and urinary vasopressin in normal subjects

We investigated the effects of sodium depletion and orthostasis on the plasma concentration and urinary excretion of vasopressin (AVP) in eight normal female subjects. After 4 days on a sodium controlled diet (130 mEq/day), the subjects were placed on a low sodium diet (30 mEq/day) for 3 days and 120 mg of furosemide was administered orally on the first day of the low sodium regimen. Sodium depletion in the present study reduced body weight by 1.6 kg and increased hematocrit by 3.5%. A significant (p less than 0.05) increase in plasma AVP and a significant (p less than 0.05) decrease in 24-h urinary excretion of AVP were observed during sodium depletion. One-hour ambulation significantly increased plasma AVP in both control and sodium depleted phases (p less than 0.01). The percent change in plasma AVP tended to correlate with that in mean blood pressure in the control phase (r = 0.69, 0.05 less than p less than 0.1), and significantly correlated in the sodium depleted phase (r = 0.86, p less than 0.01). The present results suggest that AVP may play an important role in the maintenance of blood pressure during orthostasis in the sodium depleted state.     

Molecular cloning of cDNA for sarcocystatin A and analysis of the expression of the sarcocystatin A gene during development of Sarcophaga peregrina

Sarcocystatin A is a cysteine proteinase inhibitor purified from the hemolymph of Sarcophaga peregrina larvae [Suzuki, T., & Natori, S. (1985) J. Biol. Chem. 260, 5115-5120]. We isolated a cDNA clone for sarcocystatin A and analyzed the structure and expression of the sarcocystatin A gene. Sarcocystatin A consists of 102 amino acid residues. Significant homology was found between amino acid sequences of sarcocystatin A and other mammalian cystatins, and highly conserved sequences among mammalian cystatins were also found in sarcocystatin A. Using cloned cDNA as a probe, we investigated expression of the sarcocystatin A gene during the development of Sarcophaga. Results showed that this gene was transiently activated in the very early embryonic stage and in the pupal stage, suggesting that sarcocystatin A participates in morphogenesis of larval and adult structures of Sarcophaga.     

Immunohistochemical localization of proteoglycans in interstitial elements of human pancreas and biliary system

Summary The immunohistochemical localization of large proteoglycan and small proteoglycan was observed, using antibodies 2B1 and 6B6 (Sobueet al., 1988, 1989a), in fetal and adult pancreas and biliary system as well as in tumour tissues, obtained from 11 autopsies and 74 biopsies. The distribution of chondroitin 4- and 6-sulphate side chains, type I and IV collagen and elastin were also studied. In adult pancreas and all the biliary tracts examined, periductal fibrous tissues consisted mainly of dermatan sulphate small proteoglycan with networks of fibrous elements, which were composed of large proteoglycan, elastin, type I collagen and type IV collagen. In the interstitial components of cystadenoma of pancreas and biliary duct carcinoma, similar small proteoglycan-rich components were relatively abundant, although large proteoglycan was present in much larger amounts than that in non-neoplastic adult tissues. In some cholangiomas, the extra-and intracellular hyaline globules formed by the carcinoma cells were found to contain chondroitin sulphate large proteoglycan, laminin and fibronectin.The distribution of proteoglycans was observed to be different in the arterial walls of the interlobular tissues of the adult and the fetal pancreas. The biological significance of large and small proteoglycans in the interstitial connective tissues was discussed.