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J Cappello

Publications and source records attributed to J Cappello.

16 recordsLinked to original sources

Primary visual cortex and memory. Retinal position specificity and lack of size constancy at early stages of learning a visual memory task in the macaque.

Two monkeys were trained in a novel version of a delayed match-to-sample (DMS) task. They were required to fixate on a small spot at the center of the monitor and distinguish whether two gratings presented one after the other with delays up to 1.5 s in a specific visual field location were similar or not. It was found that such learning fails to transfer readily to other retinal locations. In fact, the learning was sensitive even to very small retinal displacements of the visual stimuli. Such acute retinal position specificity implies that at least a component of the learning in this particular memory task occurs at an early visual area such as the striate cortex, which has a fine-grain topographical representation. Furthermore, at early stages of learning the DMS task, when the monkeys had not generalized the learning to stimuli of different sizes, they failed to show size constancy. That is, when the display was placed at a different distance but with the same absolute size, the performance dropped. The performance was almost fully restored when, at the new display location, stimuli were changed to fit the original retinal size. This indicates that a crucial component of the learning does occur at a site even prior to size constancy. These results show that, under certain situations, an early visual area such as the primary visual cortex may be involved even in complex behaviours such as a memory task as more than just a feature-detecting area or a relay station.

Animals↗

In-situ self-assembling protein polymer gel systems for administration, delivery, and release of drugs.

Sequential block copolymers consisting of tandem repetition of amino acids have been constructed and genetically produced based on the natural repeating structures of silk and elastin protein. Combinations of silklike and elastinlike amino acid sequence blocks in a high molecular weight protein polymer are used to confer properties similar to those observed with hard block and soft block segmented polyurethanes. A certain subset of these silk-elastinlike protein compositions, termed ProLastins, will undergo an irreversible solution to gel transition in physiological, aqueous solution. The transition occurs over time and can be controlled by temperature, solution conditions, and additives which either prevent or promote hydrogen bond-mediated chain crystallization. The process involves no covalent crosslinking. Characterization of the gelling properties of various ProLastin compositions and their ability to release compounds which are incorporated directly into the gels are presented.

Amino Acid Sequence↗

Morphology and primary crystal structure of a silk-like protein polymer synthesized by genetically engineered Escherichia coli bacteria.

The morphology and primary crystal structure of SLPF, a protein polymer produced by genetically engineered Escherichia coli bacteria, were characterized. SLPF is a segmented copolymer consisting of amino acid sequence blocks modeled on the crystalline segments of silk fibroin and the cell attachment domain of human fibronectin. Wide angle x-ray scattering (WAXS), transmission electron microscopy (TEM), selected area electron diffraction (SAED), and molecular simulations were used to analyze the primary crystal structure of SLPF. TEM experiments conducted on SLPF droplets cast from formic acid on amorphous carbon film demonstrated that these protein films have a microstructure formed of woven sheaves. The sheaves are composed of well-defined whisker crystallites. The width of the whiskers, 11.8 +/- 2.2 nm, may be correlated to the length of the silk-like segment in SLPF as predicted by molecular simulations. WAXS data, TEM images, SAED, patterns, molecular simulations, and theoretical diffraction patterns all were consistent with the crankshaft model proposed for Silk I by Lotz and Keith.

Amino Acid Sequence↗

Sequence of Dictyostelium DIRS-1: an apparent retrotransposon with inverted terminal repeats and an internal circle junction sequence.

The Dictyostelium discoideum transposon DIRS-1 contains long terminal repeats that are inverted (ITRs) and nonidentical. We show here that the internal sequence contains 4158 nucleotides and encodes three open reading frames (ORFs). Two of the ORFs (ORFs 2 and 3) are colinear and overlap for more than 2000 bases. Unusual sequence conservation between the two DIRS-1 elements in the overlap region is discussed. The conserved reading frame (ORF3) contains a 200 amino acid region that bears significant homology to retrovirus reverse transcriptase. Based on this homology, we classify DIRS-1 as a possible retrotransposon and propose a model by which the nearly genomic length 4.5 kb DIRS-1 RNA could be used to generate a genomic DNA copy of DIRS-1 with nonidentical inverted terminal repeats.

Base Sequence↗

Dictyostelium transposable element DIRS-1 has 350-base-pair inverted terminal repeats that contain a heat shock promoter.

DIRS-1 is a 4.7-kilobase-pair repetitive and apparently transposable Dictyostelium genetic element that is transcribed during differentiation or after heat shock. The terminal regions of DIRS-1 are inverted repeats of 330 base pairs. The repeats are highly conserved both within a given element as well as between different members of the family (less than 10% divergence). At the distal end of all left repeats is a 32-nucleotide sequence composed almost entirely of A and T residues. In addition to this 32-base A + T sequence, the distal region of all right repeats is extended by a 28-base-pair A + T-rich sequence that is identical in all copies. The sequences flanking each DIRS-1 sequence are completely dissimilar, and there appears to be no duplication of the genomic DNA sequence at the presumed point of DIRS-1 insertion. The terminal repeats can also be found interspersed in the genome independently of the complete element. In addition, the terminal repeats carry a 15-nucleotide sequence that greatly resembles the Drosophila consensus heat shock promoter and may be involved in the transcriptional induction of the DIRS-1 sequences.

Base Sequence↗

Dictyostelium transposable element DIRS-1 preferentially inserts into DIRS-1 sequences.

Sequence analysis of genomic clones containing the intact Dictyostelium transposable element DIRS-1 reveals that in five of six cases DIRS-1 has inserted into other DIRS-1 sequences. The nucleotide sequences just beyond the endpoints of the terminal repeats of five different genomic clones can be aligned with different regions of the internal nucleotide sequence of DIRS-1. In the three genomic clones which contain flanking sequences on both sides of the element, both flanking sequences are homologous with DIRS-1. In one of these clones, both extended flanking sequences represent the full 4.1-kilobase EcoRI fragment of DIRS-1, which has been interrupted by the insertion of an intact DIRS-1 element. There is no duplication or deletion (except possibly 1 base) of the DIRS-1 sequence upon insertion of a second DIRS-1 transposon. DIRS-1-into-DIRS-1 insertions can occur in either a colinear or inverted orientation with respect to the target sequence; the target sequence need not be an intact DIRS-1 element. We also describe a cDNA clone which could be derived by transcription of a sequence that resulted from a DIRS-1-into-DIRS-1 insertion and discuss its significance concerning the function of the heat-shock promoters found in the terminal repeats of DIRS-1 and in other DIRS-1-related sequences.

Base Sequence↗

Transcription of Dictyostelium discoideum transposable element DIRS-1.

DIRS-1 is a Dictyostelium discoideum transposable element that contains heat shock promoter sequences in the inverted terminal repeats. We showed that transcription of a 4.5-kilobase polyadenylated RNA initiates at a discrete site within the left-terminal repeat of DIRS-1, downstream from heat shock promoter and TATA box sequences. This RNA represents a full-length transcript of DIRS-1. We describe a cDNA clone that contains the 4.1 kilobases of internal sequence of DIRS-1, a cDNA clone that spans the junction between the internal sequences and the right-terminal repeat, and a cDNA clone that appears to have been transcribed from a rearranged genomic copy of DIRS-1. A second DIRS-1 RNA, named E1, is transcribed on the opposite strand of DIRS-1 from the 4.5-kilobase RNA and is under control of the heat shock promoter in the right-terminal repeat. E1 transcription initiates at multiple positions both within and downstream from the right-terminal repeat. The same transcriptional initiation sites are used during normal development and during heat shock, suggesting that in all cases DIRS-1 transcription is regulated by the heat shock promoters contained within the two terminal repeats.

Base Sequence↗

Repetitive Dictyostelium heat-shock promotor functions in Saccharomyces cerevisiae.

The Dictyostelium genome contains 40 copies of a 4.7-kilobase repetitive and apparently transposable DNA sequence (DIRS-1) and about 250 smaller elements that appear to be deletions or rearrangements of DIRS-1. Transcripts of these sequences are induced during differentiation and also by heat shock treatment of growing cells. We showed that one such cloned element, pB41.6 (2.5 kilobases) contains a nucleotide sequence identical to the Drosophila consensus heat shock promotor. To test whether this sequence might indeed control the expression of DIRS-1-related RNAs, we have cloned this genomic segment into yeast cells. In yeast cells, 41.6 directs synthesis of a 1.7-kilobase RNA that is induced at least 10-fold by heat shock. Transcription initiates at about 124 bases 3' of the putative promotor sequence and terminates within the 41.6 insert. A 381-base-pair subclone that contains the putative promotor sequence is sufficient to induce the heat shock response of 41.6 in yeast cells.

Dictyostelium↗

A repetitive Dictyostelium gene family that is induced during differentiation and by heat shock.

Clone pB41-6 (2.5 kb) contains sequences that are repeated 200-300 times in the Dictyostelium genome; about 40 of these sequences are part of a 4.5 kb repeated and apparently transposable genomic element. Clone pB41-6 hybridizes to a large number of cytoplasmic polyadenylated RNAs whose accumulation begins in the first hour of differentiation. In order to understand the regulation of these repeated sequences, we have sequenced pB41-6. It contains three long open reading frames in the "sense" strand. Remarkably, about 70 bases upstream of the transcription initiation site is a sequence identical to that responsible for induction of the Drosophila heat shock genes. A search of published sequences also generated a similar sequence upstream of one of the Dictyostelium actin genes. Indeed, we found that both pB41-6-related RNAs and actin mRNAs are increased as a result of heat shocking growing cells, and that transcription of pB41-6 sequences is induced by heat shock. Thus Dictyostelium contains a set of genes that are induced as a response to heat shock or to the stresses that trigger the initiation of development. We show here that the principal component of this "stress" is not amino acid starvation but the high density of the cells.

Actins↗

Construction of recombinant plasmids containing Xenopus immunoglobulin heavy chain DNA sequences.

A recombinant cDNA plasmid containing Xenopus immunoglobulin heavy chain sequence has been constructed from Xenopus spleen poly(A)-containing RNA. The plasmid was identified by colony hybridization and a hybridization-translation assay and its identity was confirmed by DNA sequence analysis. The portion of the heavy chain sequence contained in the plasmid is 35% homologous to mammalian mu and gamma sequences. The mRNA corresponding to this plasmid is 2.5 kilobases, in close agreement with the size of mouse mu mRNA. RNA sequences complementary to the cloned sequence appear in embryos about 24 hr after fertilization, which corresponds to 24 hr before the first detectable immunoglobulin.

Animals↗

Genetic engineering of structural protein polymers.

Genetic and protein engineering are components of a new polymer chemistry that provide the tools for producing macromolecular polyamide copolymers of diversity and precision far beyond the current capabilities of synthetic polymer chemistry. The genetic machinery allows molecular control of chemical and physical chain properties. Nature utilizes this control to formulate protein polymers into materials with extraordinary mechanical properties, such as the strength and toughness of silk and the elasticity and resilience of mammalian elastin. The properties of these materials have been attributed to the presence of short repeating oligopeptide sequences contained in the proteins, fibroin, and elastin. We have produced homoblock protein polymers consisting exclusively of silk-like crystalline blocks and elastin-like flexible blocks. We have demonstrated that each homoblock polymer as produced by microbial fermentation exhibits measurable properties of crystallinity and elasticity. Additionally, we have produced alternating block copolymers of various amounts of silk-like and elastin-like blocks, ranging from a ratio of 1:4 to 2:1, respectively. The crystallinity of each copolymer varies with the amount of crystalline block interruptions. The production of fiber materials with custom-engineered mechanical properties is a potential outcome of this technology.

Amino Acid Sequence↗