Expression of threonine synthase from Solanum tuberosum L. is not metabolically regulated by photosynthesis-related signals or by nitrogenous compounds.

Although the control of carbon fixation and nitrogen assimilation has been studied in detail, little is known about the regulation of carbon and nitrogen flow into amino acids. In this paper the isolation of a cDNA encoding threonine synthase is reported (TS; EC 4.2.99.2) from a leaf lambda ZAP II-library of Solanum tuberosum L. and the transcriptional regulation of the respective gene expression in response to metabolic changes. The pattern of expression of TS by feeding experiments of detached petioles revealed that TS expression is regulated neither by photosynthesis-related metabolites nor by nitrogenous compounds. The present study suggests that the regulation of the conversion of aspartate to threonine is not controlled at the transcript level of TS. The nucleotide and deduced amino acid sequences of potato TS show homology to other known sequences from Arabidopsis thaliana and microorganisms. TS is present as a low copy gene in the genome of potato as demonstrated in Southern blot analysis. When cloned into a bacterial expression vector, the cDNA did functionally complement the Escherichia coli mutant strain Gif41. TS transcript was found in all tissues of potato and was most abundant in flowers and source leaves.

Microsurgical reconstruction of the lower extremity using the 3M microvascular coupling device in venous anastomoses.

Microsurgical reconstruction of the lower extremity presents a difficult problem to plastic surgeons; the rate of failure is higher than any other anatomical site. We reviewed our recent experience with lower extremity microsurgical reconstruction using the 3M vascular coupling device. We believe the excellent patency rate of the coupler may minimize the well-described problem of venous thrombosis in this challenging group of patients. This study involves a consecutive series of 11 patients who presented for reconstructive microsurgery of the lower extremity at NYU Medical Center hospitals between June 1 and September 1, 1994. Ten of 11 patients had free flap transfer to traumatic lower extremity injuries, whereas the remaining reconstruction was in a diabetic individual with a chronic wound. Fifteen microvascular venous anastomoses were performed; all but 1 was performed using the 3M coupler. Our experience with 11 patients, involving 14 mechanically coupled venous anastomoses, demonstrated successful use of the coupler. No intraoperative or postoperative vascular complications occurred. The overall success rate of the 3M coupler for venous anastomoses was 100%, and all microvascular free flaps were successful. We recommend using the 3M coupling device for venous anastomoses during reconstructive microsurgery of the lower extremity. Our series demonstrates the safety and effectiveness of the 3M coupler in this challenging group of patients. In addition, a secondary benefit of the 3M coupler is a significant reduction in operative time.

Isolated sigmoid tuberculosis. Report of a case.

PURPOSE: To heighten awareness of colonic tuberculosis (TB) as a once rare disease that is undergoing a resurgence in the United States. METHODS: Report of a case of isolated sigmoid tuberculosis with a brief literature review of the topic. RESULTS: TB can no longer be considered a rare disease in the United States because, in part, of the acquired immunodeficiency syndrome epidemic and because, in part, of increased immigration and lack of containment. The signs and symptoms of colonic TB are nonspecific; therefore, a high index of suspicion must be maintained. Only 20 percent of patients will have associated active pulmonary TB. Colonoscopy with multiple biopsies at ulcer margins should be performed for diagnosis. Tissue should be sent for routine histology and culture and smeared for direct visualization of acid-fast bacilli. If colonic TB is suspected, empiric treatment is warranted, despite negative histology, smear, and culture results. Patients will usually show a dramatic response in one to two weeks. Treatment is solely medical, and all patients should receive a full course of antituberculous chemotherapy. Exploratory laparotomy is necessary if diagnosis is in doubt, when there is concern about a neoplasm, or for complications including perforation, obstruction, hemorrhage, or fistulization. CONCLUSION: An increased awareness of intestinal TB coupled with familiarity of the pathophysiology, diagnostic methods, and treatment should increase the number of cases correctly diagnosed preoperatively and, therefore, improve the outcome of patients with this disease.

A second L-type isozyme of potato glucan phosphorylase: cloning, antisense inhibition and expression analysis.

In potato tubers two starch phosphorylase isozymes, types L and H, have been described and are believed to be responsible for the complete starch breakdown in this tissue. Type L has been localized in amyloplasts, whereas type H is located within the cytosol. In order to investigate whether the same isozymes are also present in potato leaf tissue a cDNA expression library from potato leaves was screened using a monoclonal antibody recognizing both isozyme forms. Besides the already described tuber L-type isozyme a cDNA clone encoding a second L-type isozyme was isolated. The 3171 nucleotide long cDNA clone contains an uninterrupted open reading frame of 2922 nucleotides which encodes a polypeptide of 974 amino acids. Sequence comparison between both L-type isozymes on the amino acid level showed that the polypeptides are highly homologous to each other, reaching 81-84% identity over most parts of the polypeptide. However the regions containing the transit peptide (amino acids 1-81) and the insertion sequence (amino acids 463-570) are highly diverse, reaching identities of only 22.0% and 29.0% respectively. Northern analysis revealed that both forms are differentially expressed. The steady-state mRNA levels of the tuber L-type isozyme accumulates strongly in potato tubers and only weakly in leaf tissues, whereas the mRNA of the leaf L-type isozyme accumulates in both tissues to the same extent. Constitutive expression of an antisense RNA specific for the leaf L-type gene resulted in a strong reduction of starch phosphorylase L-type activity in leaf tissue, but had only sparse effects in potato tuber tissues. Determination of the leaf starch content revealed that antisense repression of the starch phosphorylase activity has no significant influence on starch accumulation in leaves of transgenic potato plants. This result indicated that different L-type genes are responsible for the starch phosphorylase activity in different tissues, but the function of the different enzymes remains unclear.

Globin evolution in the genus Xenopus: comparative analysis of cDNAs coding for adult globin polypeptides of Xenopus borealis and Xenopus tropicalis.

Globin mRNAs of Xenopus borealis and Xenopus tropicalis have been cloned and sequenced. The nucleotide and derived amino acid sequences were compared with each other and with already available data from Xenopus laevis. This analysis rendered clear evidence that the common ancestor of X. laevis and X. borealis, but not of X. tropicalis, had lost one amino acid of the beta-globins prior to a genome duplication event that preceded the segregation of the former two species. Replacement-site substitutions were used to calculate a rough time scale of genome duplication and species segregation. The results suggest an ancient separation between the X. laevis and the X. tropicalis groups occurring approximately 110-120 million years ago. Analysis of the amino acid chains demonstrated various alterations. However, some functional domains, like heme-binding sites and alpha 1 beta 2 contact sites, were subject to a high degree of conservation, indicating the existence of functional constraints on them also in the genus Xenopus.