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Biomedical subjects

A S Reddy

Publications and source records attributed to A S Reddy.

At least 55 records · Page 3Linked to original sources
PubMed1996

Expression of a cyanobacterial delta 6-desaturase gene results in gamma-linolenic acid production in transgenic plants.

Gamma-linolenic acid (GLA), a nutritionally important fatty acid in human and animal diets, is not produced in oil seed crops. Many oil seed plants, however, produce significant quantities of linoleic acid, a fatty acid that could be converted to GLA by the enzyme delta 6-desaturase if it were present. As a first step to producing GLA in oil seed crops, we have cloned a cyanobacterial delta 6-desaturase gene. Expression of this gene in transgenic tobacco resulted in GLA accumulation. Octadecatetraenoic acid, a highly unsaturated, industrially important fatty acid, was also found in transgenic tobacco plants expressing the cyanobacterial delta 6-desaturase. This is the first example of engineering the production of 'novel' polyunsaturated fatty acids in transgenic plants.

Animals↗
PubMed1996

A plant kinesin heavy chain-like protein is a calmodulin-binding protein.

Calmodulin, a calcium modulated protein, regulates the activity of several proteins that control cellular functions. A cDNA encoding a unique calmodulin-binding protein, PKCBP, was isolated from a potato expression library using protein-protein interaction based screening. The cDNA encoded protein bound to biotinylated calmodulin and 35S-labeled calmodulin in the presence of calcium and failed to bind in the presence of EGTA, a calcium chelator. The deduced amino acid sequence of the PKCBP has a domain of about 340 amino acids in the C-terminus that showed significant sequence similarity with the kinesin heavy chain motor domain and contained conserved ATP- and microtubule-binding sites present in the motor domain of all known kinesin heavy chains. Outside the motor domain, the PKCBP showed no sequence similarity with any of the known kinesins, but contained a globular domain in the N-terminus and a putative coiled-coil region in the middle. The calmodulin-binding region was mapped to a stretch of 64 amino acid residues in the C-terminus region of the protein. The gene is differentially expressed with the highest expression in apical buds. A homolog of PKCBP from Arabidopsis (AKCBP) showed identical structural organization indicating that kinesin heavy chains that bind to calmodulin are likely to exist in other plants. This paper presents evidence that the motor domain has microtubule stimulated ATPase activity and binds to microtubules in a nucleotide-dependent manner. The kinesin heavy chain-like calmodulin-binding protein is a new member of the kinesin superfamily as none of the known kinesin heavy chains contain a calmodulin-binding domain. The presence of a calmodulin-binding motif and a motor domain in a single polypeptide suggests regulation of kinesin heavy chain driven motor function(s) by calcium and calmodulin.

Adenosine Triphosphatases↗
PubMed1996

Structure and expression of a plant U1 snRNP 70K gene: alternative splicing of U1 snRNP 70K pre-mRNAs produces two different transcripts.

The product of the U1 small nuclear ribonucleoprotein particle (U1 snRNP) 70K (U1-70K) gene, a U1 snRNP-specific protein, has been implicated in basic as well as alternative splicing of pre-mRNAs in animals. Here, we report the isolation of full-length cDNAs and the corresponding genomic clone encoding a U1-70K protein from a plant system. The Arabidopsis U1-70K protein is encoded by a single gene, which is located on chromosome 3. Several lines of evidence indicate that two distinct transcripts (short and long) are produced from the same gene by alternative splicing of the U1-70K pre-mRNA. The alternative splicing involves inclusion or exclusion of a region (910 bp) that we named "included intron." Two transcripts were clearly detectable in all tissues tested, and the level of the transcripts varied in different organs. The deduced amino acid (427 residues) sequence from the short transcript has strong homology to the animal U1-70K protein and contains an RNA recognition motif, a glycine hinge, and an arginine-rich region characteristic of the animal U1-70K protein. The long transcript has an in-frame translational termination codon within the 910-bp included intron, resulting in a truncated protein containing only 204 amino acids. The protein encoded by the short transcript is recognized by U1 RNP-specific monoclonal antibodies and binds specifically to the Arabidopsis U1 snRNA, whereas the protein from the long transcript does not. In addition, multiple polyadenylation sites were observed in the 3' untranslated region. These results suggest a complex post-transcriptional regulation of Arabidopsis U1-70K gene expression.

Alternative Splicing↗
PubMed1996

Complex arrangement of dispersed repeated DNA sequences in Oryza officinalis.

A 525-bp BglII fragment was isolated from Oryza officinalis DNA (accession W1278) and shown to correspond to a new dispersed repetitive DNA sequence with specificity restricted to a subset of the wild rice with a C genome. The sequence of the fragment was determined but it does not correspond to any sequence already present in databases. It contains several imperfect palindromes. Larger genomic clones (12-18 kbp) were isolated and all contain sequences homologous to the BglII element. Analysis of these clones confirms that the BglII element is dispersed in the O. officinalis genome. From one genomic clone, the sequences adjacent to the BglII element were subcloned and used as probes to demonstrate that the sequences flanking the BglII element are variable in different genomic clones and that some of them are also dispersed repetitive sequences. The genomic specificity of two of these dispersed repeats was evaluated and shown to be different from that of the initial BglII element. This analysis revealed a complex arrangement of various dispersed repeated sequences.

Bacterial Proteins↗
PubMed1995

The flexor digitorum longus: an anatomic and microscopic study for use as a tendon graft.

Recent experimental studies have shown that intrasynovial donor tendons have the capacity to heal with minimal alteration in the gliding surface when grafted to an intrasynovial location. In order to evaluate the flexor digitorum longus of the foot as a potential matched donor tendon for use in flexor tendon grafting, we dissected 15 feet (75 toes) and 10 hands (50 fingers) to determine the intrasynovial lengths of the flexor digitorum longus and flexor digitorum profundus and to determine a method of tendon graft harvest. Specimens were taken from the intrasynovial portion of each tendon for evaluation by light and electron microscopy. The intrasynovial tendons of the hand and foot were similar in vascularity, cellularity, gliding surface, and ultrastructure. The flexor digitorum longus tendon of the foot was a mean length of 12 cm to the area of tendon confluence in the midfoot, which will usually provide enough tendon length for grafting from the distal phalanx to the palm of the hand. Because of the difference in length of the synovial sheaths of the fingers and toes, some extrasynovial tendon will be transferred to the synovial space when a flexor digitorum longus tendon is used.

Cadaver↗
PubMed1995

Characterization of a dispersed repetitive DNA sequence associated with the CCDD genome of wild rice.

A HindII repetitive fragment (pOD3) was isolated and cloned from the genomic DNA of an accession of Oryza latifolia, a wild rice species that possesses a tetraploid CCDD genome. Southern blot analysis using this clone as a probe demonstrated that this repetitive DNA sequence had a dispersed organization in the CCDD genome and seemed to be highly specific for this genome type. This fragment is the first CCDD-specific repeated DNA sequence to be described. The hybridization pattern is similar for most CCDD accessions tested, although a few showed no hybridization signal. The nucleotide sequence of the element cloned in pOD3 was determined and analysed. The 1783 base pair long repeated sequence shows no homology with other known nucleotide sequences. In addition, none of the amino acid sequences deduced from the potential open reading frames contained in the pOD3 repeat is homologous to any known protein. The nucleotide sequence presents several internal repeats, direct or inverted, but their significance remains unknown.

Base Sequence↗
PubMed1994

Cloning of a family of cyclins from Arabidopsis thaliana.

Degenerate oligonucleotide primers corresponding to two conserved regions in animal cyclins were used to amplify cyclin sequences from Arabidopsis thaliana by polymerase chain reaction (PCR). Screening of a floral meristem cDNA library with two distinct PCR-generated cyclin sequences resulted in the isolation of four different cyclin cDNAs. Southern analysis of genomic DNA and sequence information from the isolated clones suggest that there is a family of cyclins in Arabidopsis.

Amino Acid Sequence↗
PubMed1994

Calcium-dependent proteinase activity in root cultures of Arabidopsis.

The presence of calcium-dependent proteinase in plants was investigated using soluble proteins from Arabidopsis root cultures. Calcium increased the proteolytic activity by two to three fold. Calcium-stimulated proteolytic activity was inhibited by different protease inhibitors. The highest inhibition (79%) of calcium-dependent proteolytic activity was observed in the presence of leupeptin. Thirty-eight percent of calcium-dependent proteolytic activity was inhibited by E-64, an inhibitor of cysteine proteinases. Calmodulin did not have any effect on calcium-dependent as well as calcium-independent proteolytic activity in the cellular extract. These results provide the first evidence for the presence of calcium activated proteinase in plants and open new avenues to investigate the mode of calcium action in plants.

Arabidopsis↗
PubMed1994

Developmental and hormonal regulation of sunflower helianthinin genes: proximal promoter sequences confer regionalized seed expression.

DNA elements involved in the regulation of two sunflower helianthinin genes were identified by analysis of beta-glucuronidase (GUS) expression in transgenic tobacco driven by sequences derived from the 5' upstream regions of these genes. A 2.4-kb upstream region of the helianthinin gene HaG3-A conferred rigorous developmental GUS expression in transgenic tobacco seeds with no significant GUS activity in nonembryonic tissues. Regions of the helianthinin upstream regulatory ensemble (URE) conferred ectopic expression in nonembryonic tissues when analyzed outside of the context of the complete helianthinin regulatory complex. A proximal promoter region was identified that conferred significant GUS expression in seeds but not in leaves of transgenic tobacco. Three sequence motifs that bind to seed nuclear proteins were identified in the proximal promoter region; mutations in these motifs significantly reduced the level of nuclear protein binding. Another important class of cis-regulatory elements was identified in the helianthinin URE that conferred abscisic acid-responsive GUS expression. In the full-length helianthinin URE, these elements only responded to abscisic acid in the developing seed, suggesting that the helianthinin gene contains additional regulatory elements, possibly in the proximal promoter region, that ensure hierarchical control in the developing seed.

2S Albumins, Plant↗
PubMed1993

Isolation of a delta 6-desaturase gene from the cyanobacterium Synechocystis sp. strain PCC 6803 by gain-of-function expression in Anabaena sp. strain PCC 7120.

The enzyme delta 6-desaturase is responsible for the conversion of linoleic acid (18:2) to gamma-linolenic acid (18:3 gamma). A cyanobacterial gene encoding delta 6-desaturase was cloned by expression of a Synechocystis genomic cosmid library in Anabaena, a cyanobacterium lacking delta 6-desaturase. Expression of the Synechocystis delta 6-desaturase gene in Anabaena resulted in the accumulation of gamma-linolenic acid (GLA) and octadecatetraenoic acid (18:4). The predicted 359 amino acid sequence of the Synechocystis delta 6-desaturase shares limited, but significant, sequence similarity with two other reported desaturases. Analysis of three overlapping cosmids revealed a delta 12-desaturase gene linked to the delta 6-desaturase gene. Expression of Synechocystis delta 6- and delta 12-desaturases in Synechococcus, a cyanobacterium deficient in both desaturases, resulted in the production of linoleic acid and gamma-linolenic acid.

Amino Acid Sequence↗
PubMed1993

Calcium and signal transduction in plants.

Environmental and hormonal signals control diverse physiological processes in plants. The mechanisms by which plant cells perceive and transduce these signals are poorly understood. Understanding biochemical and molecular events involved in signal transduction pathways has become one of the most active areas of plant research. Research during the last 15 years has established that Ca2+ acts as a messenger in transducing external signals. The evidence in support of Ca2+ as a messenger is unequivocal and fulfills all the requirements of a messenger. The role of Ca2+ becomes even more important because it is the only messenger known so far in plants. Since our last review on the Ca2+ messenger system in 1987, there has been tremendous progress in elucidating various aspects of Ca(2+) -signaling pathways in plants. These include demonstration of signal-induced changes in cytosolic Ca2+, calmodulin and calmodulin-like proteins, identification of different Ca2+ channels, characterization of Ca(2+) -dependent protein kinases (CDPKs) both at the biochemical and molecular levels, evidence for the presence of calmodulin-dependent protein kinases, and increased evidence in support of the role of inositol phospholipids in the Ca(2+) -signaling system. Despite the progress in Ca2+ research in plants, it is still in its infancy and much more needs to be done to understand the precise mechanisms by which Ca2+ regulates a wide variety of physiological processes. The purpose of this review is to summarize some of these recent developments in Ca2+ research as it relates to signal transduction in plants.

Amino Acid Sequence↗
PubMed1993

Characterization of new variants of a satellite DNA from Oryza officinalis, specific for the CC genome of wild rice.

Two groups of tandemly repeated DNA fragments have been isolated and cloned from the Oryza officinalis genome (W 1278). These fragments have been sequenced and are 374 and 367 bp long. They were compared with an element previously isolated from another O. officinalis accession. They are 76 and 92% homologous to the latter and show 78% homology between themselves. The two types of elements are interspersed within large blocks of tandemly organized units. Their copy number is in the range of 200,000 units. This sequence was present only in wild rice accessions with a CC genome. However, this apparent specificity is even more restricted, since it was absent from some O. officinalis accessions and from the allotetraploid species of the CCDD type. Use of digestions with 4-bp restriction enzyme cutters and analysis on denaturing polyacrylamide gels revealed polymorphism that might be used to map the repeats in the genome as well as to establish the relationship between accessions. Finally, hybridization of the repeated sequence to DNA prepared from the eight available O. sativa-O. officinalis monosomic addition lines revealed that the elements are located on two alien chromosomes. This suggests that these elements have at least some restricted chromosome specificity.

Base Sequence↗
PubMed1993

Restriction of the injury response following an acute muscle strain.

Indirect skeletal muscle injuries have been found to occur exclusively near the myotendinous junction (MTJ). Although muscle cells are syncytial and extend far into the muscle belly, the response to this injury has been shown to be limited to a focal area near the MTJ. This study examined single muscle fibers near their site of rupture in order to document structural changes that may help to explain this limited injury response. A partial, nondisruptive strain injury was created in a New Zealand white rabbit extensor digitorum longus muscle. The muscles were left in vivo for 60 min or 6 h before harvesting. The specimens were divided into four groups of single fibers: ruptured fibers (60 min) attached to muscle belly (group I); ruptured fibers (60 min) attached to tendon (group II); ruptured fibers (6 h) attached to muscle belly (group III); and normal, unstretched fibers (60 min) at the MTJ (group IV) (N = 10 for each group). Sarcomeres closest to the site of fiber rupture in the three injured groups (I, II, and III) were hypercontracted, with lengths well below the physiologic range (I: 0.86 +/- 0.08 microns, II: 0.96 +/- 0.09 microns, III: 0.96 +/- 0.21 microns). There was a progressive increase in sarcomere length, which became normal by 300-500 microns away from the site of rupture (I: 2.17 +/- 0.48 microns, II: 2.69 +/- 0.42 microns, III: 2.08 +/- 0.48 microns). The 6-h fibers in group III showed evidence of necrosis before the transition to normal sarcomere spacing occurred.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Cloning of the cDNA for U1 small nuclear ribonucleoprotein particle 70K protein from Arabidopsis thaliana.

We cloned and sequenced a plant cDNA that encodes U1 small nuclear ribonucleoprotein (snRNP) 70K protein. The plant U1 snRNP 70K protein cDNA is not full length and lacks the coding region for 68 amino acids in the amino-terminal region as compared to human U1 snRNP 70K protein. Comparison of the deduced amino acid sequence of the plant U1 snRNP 70K protein with the amino acid sequence of animal and yeast U1 snRNP 70K protein showed a high degree of homology. The plant U1 snRNP 70K protein is more closely related to the human counter part than to the yeast 70K protein. The carboxy-terminal half is less well conserved but, like the vertebrate 70K proteins, is rich in charged amino acids. Northern analysis with the RNA isolated from different parts of the plant indicates that the snRNP 70K gene is expressed in all of the parts tested. Southern blotting of genomic DNA using the cDNA indicates that the U1 snRNP 70K protein is coded by a single gene.

Amino Acid Sequence↗
PubMed1990

Molecular cloning of cDNAs for auxin-induced mRNAs and developmental expression of the auxin-inducible genes.

By differential hybridization, two auxin-inducible cDNA clones (lambda SAR1 and lambda SAR2) have been isolated from a cDNA library constructed to poly(A)+ mRNA from auxin-treated strawberry receptacles. Both the clones have been used as probes to study the expression of the auxin-induced genes in pollinated and unpollinated fruits of various stages of development and in different organs. A high level of auxin-induced mRNAs is found in pollinated fruits as compared to unpollinated fruits of the same age, suggesting that the expression of the auxin-induced genes is developmentally regulated and the level of auxin-induced mRNAs is regulated by endogenous auxin. Furthermore, our data on the expression of lambda SAR1 and lambda SAR2 genes in pollinated and unpollinated fruits revealed a positive correlation between growth of strawberry fruit and the induction of mRNA corresponding to the lambda SAR1 and lambda SAR2 clones. Ethylene has no effect on the expression of the auxin-induced mRNAs. lambda SAR1 mRNA is not detected in other parts of strawberry plants whereas lambda SAR2 mRNA is present in roots. Furthermore, mRNA corresponding to lambda SAR1 and lambda SAR2 is not detected in other auxin-responsive plant systems such as pea epicotyls and bean explants.

Cloning, Molecular↗
PubMed1990

Molecular cloning and sequencing of a cDNA for an auxin-repressed mRNA: correlation between fruit growth and repression of the auxin-regulated gene.

A complementary DNA (cDNA) library has been constructed in lambda gt 10 from poly(A)+ mRNA isolated from auxin-deprived strawberry receptacles. By differential plaque filter hybridization, a cDNA (lambda SAR5) to an auxin-repressed mRNA has been isolated. The expression of the auxin-repressed gene is studied at various stages of normal fruit development and in fruits of variant strawberry genotype using lambda SAR5 as a probe. Northern analyses of RNA isolated from pollinated and unpollinated fruits of various developmental stages revealed that mRNA corresponding to the lambda SAR5 clone is repressed during normal fruit development, and the level of lambda SAR5 mRNA is regulated by endogenous auxin. Furthermore, results with both normal and variant genotype strawberry fruit indicate that there is a positive correlation between growth of strawberry fruit and repression of mRNA corresponding to the lambda SAR5 clone. The lambda SAR5 cDNA has been sequenced and is 723 nucleotides in length. The deduced protein has 111 amino acid residues with a molecular mass of 12.5 kDa. The putative polypeptide starts at nucleotide position 20 and ends at 352. The molecular weight of the predicted polypeptide is in agreement with the molecular weight of the in vitro translated polypeptide of hybrid selected mRNA. A comparison of the nucleotide and deduced amino acid sequence of lambda SAR5 with nucleotide and protein sequences in data banks has not revealed any homology to known proteins.

Amino Acid Sequence↗