A genome survey indicates a possible susceptibility locus for bipolar disorder on chromosome 22.

Bipolar disorder or manic depressive illness is a major psychiatric disorder that is characterized by fluctuation between two abnormal mood states. Mania is accompanied by symptoms of euphoria, irritability, or excitation, whereas depression is associated with low mood and decreased motivation and energy. The etiology is currently unknown; however, numerous family, twin, and adoption studies have argued for a substantial genetic contribution. We have conducted a genome survey of bipolar disorder using 443 microsatellite markers in a set of 20 families from the general North American population to identify possible susceptibility loci. A maximum logarithm of odds score of 3.8 was obtained at D22S278 on 22q. Positive scores were found spanning a region of nearly 32 centimorgans (cM) on 22q, with a possible secondary peak at D22S419. Six other chromosomal regions yielded suggestive evidence for linkage: 3p21, 3q27, 5p15, 10q, 13q31-q34, and 21q22. The regions on 22q, 13q, and 10q have been implicated in studies of schizophrenia, suggesting the possible presence of susceptibility genes common to both disorders.

Induction of swelling, synovial hyperplasia and cartilage proteoglycan loss upon intra-articular injection of transforming growth factor beta-2 in the rabbit.

Transforming growth factor beta (TGF-beta) is a multifunctional homodimeric polypeptide with potent actions upon many target cells, including those of mesenchymal and haemopoietic lineage. The recent reports of high levels of the cytokine in rheumatoid synovium and synovial fluid, prompted this study into the effect of intra-articular injection of TGF beta-2 into rabbit knee-joints. Four daily injections of 1 microgram caused swelling, probably as a consequence of prostaglandin E2 production, synovial fibroblastic hyperplasia and a striking loss of femoral condyle proteoglycan. Using the polymerase chain reaction, no evidence could be obtained for the induction of interleukin-1 alpha gene expression in either synovial tissue or synovial fluid cells. These findings suggest that the TGF-beta present in the rheumatoid joint may contribute directly to the pathogenesis of rheumatoid arthritis.

Specific binding of transposase to terminal inverted repeats of transposable element Tn3.

Tn3 transposase, which is required for transposition of Tn3, has been purified by a low-ionic-strength-precipitation method. Using a nitrocellulose filter binding assay, we have shown that transposase binds to any restriction fragment. However, binding of the transposase to specific fragments containing the terminal inverted repeat sequences of Tn3 can be demonstrated by treatment of transposase-DNA complexes with heparin, which effectively removes the transposase bound to the other nonspecific fragments at pH 5-6. DNase I "footprinting" analysis showed that the transposase protects an inner 25-base-pair region of the 38-base-pair terminal inverted repeat sequence of Tn3. This protection is not dependent on pH. Interestingly, binding of the transposase to the inverted repeat sequences facilitates DNase I to nick at the end of the Tn3 sequence. It was also observed that the transposase protects the end regions of restriction fragments with a cohesive sequence at their 5' end or with a flush end from DNase I cleavage. The specific and nonspecific binding of transposase to DNA is ATP-independent.

Identification of a repeated sequence element required for efficient encapsidation of the adenovirus type 5 chromosome.

Adenovirus type 5 deletion mutants that lack portions of their cis-acting DNA encapsidation signal synthesized nearly normal levels of viral DNA and late polypeptides but failed to efficiently package the DNA into virus particles. A series of mutant viruses carrying small deletions were produced and used to identify a repeated element (AGTAAATTTGGGC and AGTAAGATTTGGCC) as a key component of the packaging signal. One copy of the repeat was sufficient to signal efficient packaging. The packaging domain could function near either end of the viral chromosome but was no longer active when moved several hundred base pairs toward the interior of the DNA molecule.

ATP-dependent specific binding of Tn3 transposase to Tn3 inverted repeats.

Transposons are discrete segments of DNA which are capable of moving from one site in a genome to many different sites. Tn3 is a prokaryotic transposon which is 4,957 base pairs (bp) long and encodes a transposase protein which is essential for transposition. We report here a simple method for purifying Tn3 transposase and demonstrate that the transposase protein binds specifically to the ends of the Tn3 transposon in an ATP-dependent manner. The transposase protein binds to linear double-stranded DNA both nonspecifically and specifically; the nonspecific DNA binding activity is sensitive to challenge with heparin. Site-specific DNA binding to the ends (inverted repeats) of Tn3 is observed only when binding is performed in the presence of ATP; this ATP-dependent site-specific DNA binding activity is resistant to heparin challenge. Our results indicate that ATP qualitatively alters the DNA binding activity of the transposase protein so that the protein is able to bind specifically to the ends of the Tn3 transposon.

Escherichia coli RNA polymerase binding sites and transcription initiation sites in the transposon Tn3.

We have identified the Escherichia coli RNA polymerase-binding sites and the transcription initiation sites in the transposon Tn3. Results from nitrocellulose filter-binding assays indicate that there are two regions within Tn3 capable of forming stable binary complexes with RNA polymerase. The two regions are a 208-bp region containing the N-terminal coding sequence of the transposase (tnpA) and repressor (tnpR) genes, and a 332-bp region containing the N-terminal coding sequence for the beta-lactamase (bla) gene. DNase I footprint analysis of the 208-bp and 332-bp fragments further defined an extended region of protection, approx. 110 bp long, located between the transposase and repressor coding regions, and an 80-bp region of protection near the N-terminal coding sequence of the beta-lactamase gene. In vitro transcription studies with fragments containing these protected regions allowed us to determine the precise transcription initiation sites for the transposase, repressor, and beta-lactamase mRNAs. The transposase and repressor mRNAs are transcribed divergently and their transcription initiation sites are separated by 80 bp. The -35 homology regions for the transposase and repressor promoters are separated by 10 bp and the -10 homology region of the transposase promoter is coincident with the recombination site (res) for the site-specific recombinase activity (resolvase) of the repressor protein, which is required for resolution of Tn3 cointegrates. We discuss the significance of this complex divergently transcribed promoter region with respect to regulation of Tn3 transposition and we propose a model for coordinated regulation of the tnpA and tnpR genes. We also compare the Tn3 tnpA-tnpR intercistronic region with that of the closely related transposon gamma delta.

Poliovirus replication proteins: RNA sequence encoding P3-1b and the sites of proteolytic processing.

A partial amino-terminal amino acid sequence of each of the major proteins encoded by the replicase region (P3) of the poliovirus genome has been determined. A comparison of this sequence information with the amino acid sequence predicted from the RNA sequence that has been determined for the 3' region of the poliovirus genome has allowed us to locate precisely the proteolytic cleavage sites at which the initial polyprotein is processed to create the poliovirus products P3-1b (NCVP1b), P3-2 (NCVP2), P3-4b (NCVP4b), and P3-7c (NCVP7c). For each of these products, as well as for the small genome-linked protein VPg, proteolytic cleavage occurs between a glutamine and a glycine residue to create the amino terminus of each protein. This result suggests that a single proteinase may be responsible for all of these cleavages. The sequence data also allow the precise positioning of the genome-linked protein VPg within the precursor P3-1b just proximal to the amino terminus of polypeptide P3-2.