The Alu I-induced bands in metaphase chromosomes of orangutan (Pongo pygmaeus)

Summary Restriction endonucleases have been recently proved to be active on fixed chromosomes, thus they are useful in chromatin structure studies. Within this class of enzymes, Alu I is able to detect the presence and localization of highly repetitive DNA sequences in human and in other mammalian and dipteran species. In this paper the pattern obtained on fixed metaphase chromosomes of orangutan (Pongo pygmaeus) by Alu I digestion and Giemsa staining is shown. The results are discussed in the light of the distribution, in this species, of the I–IV human satellite DNAs. It is also suggested that in Pongo some highly repetitive sequences, different from the major human satellites, are present.     

Procedure for mutagenizing spores of Saccharomyces cerevisiae

A procedure for inducing mutants of a homothallic strain of Saccharomyces cerevisiae is described. The essential parts of the procedure are long incubation in Glusulase, which preferentially kills vegetative cells instead of spores, and treatment in 9% ethyl methanesulfonate, which also preferentially kills vegetative cells instead of spores. Consequently, the viable population is virtually 100% spores.     

Cloning and sequencing of a full length cDNA coding for human retinol-binding protein.

We have isolated and sequenced a cDNA clone coding for human Retinol Binding Protein. The sequence indicates that Retinol Binding Protein is synthesized as a single polypeptide chain precursor which is then matured to the secreted protein by removal of a leader peptide. Southern and Northern blot analysis suggest that the gene is present in one or few copies per haploid genome and is transcribed in a single mRNA species.     

Sequence of human haptoglobin cDNA: evidence that the alpha and beta subunits are coded by the same mRNA.

We have isolated and sequenced a cDNA clone coding for human haptoglobin. Our sequence shows that haptoglobin is very likely synthesized as a single polypeptide chain which is then cleaved at an Arg residue to generate its two characteristic alpha and beta subunit. Southern blot analysis suggests that there are at least two copies of the haptoglobin gene per haploid genome.     

Neuronal nets and nerve cell interactions in vitro in insect systems

Summary The development of a synthetic medium that supports growth and differentiation of insect embryonic tissues afforded the possibility of studying the interactions between nerve and other cell types in long term cultures. The mechanical dissociation of embryonic nerve tissues results in survival of nerve cells but not of glial cells. The dissociated glial-free neurons produce a dense fibrillar network in the presence, but not in the absence, of foregut explants or other tissues from same donors. Nerve fiber bundles outgrowing from dissociated neurons enter foregut segments and establish synaptic connections with muscle cells. Foregut explants undergo differentiation and become contractile in long term cultures when innervated by dissociated nerve cells. The progressive deterioration of similar foregut tissues cultured alone contrasts with the excellent condition of innervated explants and suggests that this is due to trophic factors released by nerve fibers. The same in vitro systems provided the opportunity of studying the interaction between nerve fibers produced by the autonomic ingluvial ganglion, which adheres to the surface of the alimentary tract, and muscle cells. Multiple esophagus explants from cockroach embryos become interconnected by fibers emerging from ingluvial ganglia, when the explants are combined in vitro at short distance from each other. Muscle cells migrating from the esophagi line up on axons branching out in the medium, or form contractile ribbons which, in turn, establish connections with nerve fibers. The thigmotropism of muscle cells and strong affinity for nerve fibers reveal a new aspect of muscle cells-to-fibers interaction, amenable to further analysis in vitro. This work was supported in part by United States Public Health Service grant NS-03777 and grant GB-16330 X from the National Science Foundation     

Mechanism of Excretion of a Bacterial Proteinase: Factors Controlling Accumulation of the Extracellular Proteinase of a Sarcina Strain (Coccus P)

It has been known that the extracellular proteinase of Coccus P is found only in cultures grown in the presence of Ca²⁺. It is now shown that this cation is required neither for synthesis, excretion, or activation of a zymogen nor as a prosthetic factor necessary for enzymatic activity. The only function of Ca²⁺ is to stabilize the active structure of the enzyme molecule, presumably by substituting for absence of S-S bridges. In the absence of Ca²⁺, the excreted proteinase undergoes rapid autodigestion and, instead of the active protein, its hydrolytic products are accumulated in the culture fluid. In minimal medium and under conditions of enzyme stability [presence of Ca²⁺ and Ficoll (Pharmacia)], Coccus P accumulates the proteinase at a gradually reduced speed although the rate of cultural growth remains constant. It is shown that this decline in rate of accumulation is caused by the excreted proteinase itself, possibly acting on its own precursor emerging from the cell in a form susceptible to proteolytic attack and not amenable to Ca²⁺ protection. A proteinase precursor is actually demonstrable in a calciumless culture at the onset of the enzyme accumulation which follows Ca²⁺ addition. It is suggested that excreted proteins require an unfolded (or incompletely folded) structure to cross the cell envelope.     

Growth-associated modifications of low-molecular-weight thiols and protein sulfhydryls in human bronchial fibroblasts

The thiol redox status of cultured human bronchial fibroblasts has been characterized at various growth conditions using thiol-reactive monobromobimane, with or without the combination of dithiotreitol, a strong reducing agent. This procedure has enabled measurement of the cellular content of reduced glutathione (GSH), total glutathione equivalents, cysteine, total cysteine equivalents, protein sulfhydryls, protein disulfides, and mixed disulfides. Passage of cells with trypsin perturbs the cellular thiol homeostasis and causes a 50% decrease in the GSH content, whereas the total cysteine content is subsequently increased severalfold during cell attachment. During subsequent culture, transient severalfold increased levels of GSH, protein-bound thiols, and protein disulfides are reached, whereas the total cysteine content gradually declines. These changes in the redox balance of both low-molecular-weight thiols and protein-bound thiols correlate with cell proliferation and mostly precede the major growth phase. When the onset of proliferation is inhibited by maintenance of cells in medium containing decreased amounts of serum, the GSH content remains significantly increased. Subsequent stimulation of growth by addition of serum results in decreased GSH levels at the onset of proliferation. In thiol-depleted medium, proliferation is also inhibited, whereas GSH levels are increased to a lesser extent than in complete medium. Exposure to buthionine sulfoximine inhibits growth, prevents GSH synthesis, and results in accumulation of total cysteine, protein-bound cysteine, and protein disulfides. For extracellular cystine, variable rates of cellular uptake correlate with the initial increase in the total cysteine content observed following subculture and with the GSH peak that precedes active proliferation. The results strongly suggest that specific fluctuations in the cellular redox balance of both free low-molecular-weight thiols and protein sulfhydryls are involved in growth regulation of normal human fibroblasts.     

Le Fort fractures without mobility

The Le Fort fracture without maxillary mobility constitutes 9 percent of maxillary fractures observed over a 3-year period. A high Le Fort (level II or III) injury exists as a one- or two-piece incomplete fracture. The degree of fracture is insufficient to permit mobility of the maxillary alveolus. Frequently, an obvious unilateral zygomatic fracture is present. Physical findings consist of bilateral eyelid ecchymosis and malocclusion. The occlusal disturbance may consist of either crossbite, open bite, maxillary rotation, or lack of proper dental intercuspation. On CT scan, fractures are best demonstrated in the posterior and medial maxillary walls at the Le Fort I level; they are most obvious unilaterally with contralateral fractures that may be subtle. Bilateral maxillary sinus fluid is consistently present on CT. Treatment usually consists of observation and traction elastics but may require mobilization of the fragments followed by open reduction and rigid fixation.