Human λ light chain locus: Organization and DNA sequences of three genomicJ regions

Evidence for the genomic organization of human lambda light chain joining (J) region gene segments is presented. A mouse J probe was used in Southern hybridizations to localize joining region sequences in a cosmid clone containing the genomic cluster of six human lambda constant (C) region gene segments. The results of these hybridizations suggest the presence of at least one J gene segment upstream from each constant region gene segment. The DNA sequences indicate that the human JI, J2, and J3 gene segments have consensus nonamer and heptamer sequences, proposed to be involved in V-J joining, are capable of encoding the known amino acid sequences for the respective J peptides, and have a sequence which could give functional RNA splice site at the end of their coding regions. Our data show that a single functional J is located 1.3 or 1.6 kb upstream of each of the C gene segments known to encode the Mcg, Kern^– Oz^–, and Kern^–Oz⁺ isotypes. Therefore, the gene organization of this region of the human lambda locus is J1 CI -J2C2-J3C3. The DNA sequences ofJ 1,J 2, andJ 3 presented in this paper establish that a singleJ gene segment precedes each expressed C gene segment, and support a model for the evolution of the human JC clusters where JICI andJ2C2-J3C3. arose from different ancestral JC units.     

Operculum closing as a defence against predatory leeches in four British freshwater prosobranch snails

The defence reaction of operculum closing in response to the presence of the molluscivorous leech Glossiphonia complanata (L.) and the non-molluscivorous Erpobdella octoculata (L.) was studied in four species of freshwater prosobranch gastropod. Bithynia tentaculata (L.) and Valvata piscinalis (Müller) can distinguish between the leeches, reacting only to G. complanata. V. piscinalis is capable of a greater degree of distance chemoreception of the leech ‘scent’. Valvata cristata Müller and Potamopyrgus jenkinsi (Smith) did not react to either leech. V. cristata may not be a potential prey item for G. complanata, while P. jenkinsi is fed on by the leech, but is a relative newcomer to the freshwater fauna. Animal Ecology Research Group, Department of Zoology, University of Oxford Commonwealth Forestry Institute, Department of Plant Sciences, University of Oxford     

Influence of sequence and distance between two operators on interaction with the lac repressor

The influence of additional operator or pseudooperator sequences on the lactose repressor-operator interaction has been investigated. Results of kinetic and equilibrium binding measurements suggest an important in vivo role for the Z-gene pseudooperator in repressor-operator binding; the formation of a ternary, looped complex is indicated by the influence of secondary operator sites on binding parameters. Although the binding affinity of the Z-gene pseudooperator [Oz] is only approximately 1/30 that observed for the primary operator [O], the binding affinity to DNA containing both Oz and O is significantly higher than either sequence alone or the sum of the two. This synergistic effect is enhanced further by replacing the pseudooperator sequence [Oz] with the primary operator sequence and results in an even stronger ternary complex in plasmids with duplicate primary sites. The distance between the center of the two primary operators affects the formation of a ternary complex in the linear DNA molecules. Decreased dissociation rate constants were observed with spacing of operator-like sequences between 300 and 500 base pairs (bp). Minimal influence of the second operator on repressor binding is observed when the operators are separated by approximately 4000 or approximately 100 bp. The significant influence of distance on kinetic and equilibrium parameters was demonstrated by measurements on plasmid pRW1511 [Oi-O-PL-Oz] cleaved with restriction enzymes either in the polylinker region to place Oi-O and Oz on opposite ends of the linear plasmid or outside this region to maintain the sites within 500 bp. These results are consistent with the formation of operator-repressor-pseudooperator ternary complex to generate a looped DNA structure.     

Influence of supercoiling and sequence context on operator DNA binding with lac repressor

The dissociation of the repressor-operator complex from a series of negatively supercoiled plasmid DNAs was examined as a function of the sequence context, orientation, and spacing. The plasmids were grouped into four classes, each with common sequence context. The highest dissociation rate constants were observed for the plasmids containing only a single operator (or pseudooperator) sequence, while approximately 10-fold lower rate constants were measured for plasmids with the I gene pseudooperator in conjunction with either the Z gene pseudooperator or the primary operator. Comparison of the behavior of these two classes of plasmids demonstrated the importance of two operator sequences and supported a model of DNA loop formation to stabilize the repressor-operator complex (Whitson, P. A., and Matthews, K. S. (1986) Biochemistry 25, 3845-3852; Whitson, P. A., Olson, J. S., and Matthews, K. S. (1986) Biochemistry 25, 3852-3858; Whitson, P. A., Hsieh, W. T., Wells, R. D., and Matthews, K. S. (1987) J. Biol. Chem. 262, 4943-4946; Kr?mer, H., Niem?ller, M., Amouyal, M., Revet, B., von Wilcken-Bergmann, B., and Müller-Hill, B. (1987) EMBO J. 6, 1481-1491). The third class, with intermediate dissociation rate constants, was comprised of plasmids which contained the primary operator and the higher affinity pseudooperator normally located in the Z gene. Neither the additional presence of the I gene pseudooperator nor the orientation of the primary operator relative to the Z gene pseudooperator significantly affected the dissociation rate constants. The binding characteristics of this group of plasmids demonstrated the essential role of the Z gene pseudooperator in the formation of intramolecular ternary complex and suggested an in vivo function for this pseudooperator. Plasmids containing two primary operator sequences were the class with lowest dissociation rate constants from lac repressor, and minimal effects of salt or spacing on dissociation of this class were observed. These data are consistent with formation of an intramolecular complex with a looped DNA segment stabilized by the combination of increased local concentration of binding sites and torsional stresses on the DNA which favor binding in supercoiled DNA.     

Pancreatic B-cell function in non-insulin-dependent diabetes mellitus during successive periods of sulfonylurea and insulin treatment: serum C-peptide response to glucagon and urine C-peptide excretion

Serum C-peptide responses to glucagon and daily urine C-peptide excretion in successive periods of different treatment in two groups of patients with non-insulin-dependent diabetes mellitus (NIDDM) (mean interval between two tests less than 1 month) were compared. In group A patients (n = 8), the glycemic control was improved after transferring the treatment from sulfonylurea (SU) to insulin (fasting plasma glucose: SU: 192 +/- 47, insulin: 127 +/- 21 mg/dl, mean +/- S.D., p less than 0.01). Fasting serum C-peptide immunoreactivity (CPR) was significantly lower at the period of insulin treatment (SU: 1.93 +/- 1.01, insulin: 1.47 +/- 0.79 ng/ml, p less than 0.05), but there was no difference in the increase in serum CPR (maximal--fasting) (delta serum CPR) during glucagon stimulation in the two periods of treatment (SU: 1.70 +/- 0.72, insulin: 1.47 +/- 0.98 ng/ml). In group B patients (n = 7), there was no significant difference in glycemic control after transferring the treatment from insulin to SU (fasting plasma glucose: insulin: 127 +/- 24, SU: 103 +/- 13 mg/dl). Fasting serum CPR was significantly lower during the period of insulin treatment (insulin: 1.39 +/- 0.64, SU: 2.21 +/- 0.86 ng/ml, p less than 0.025), but delta serum CPR during glucagon stimulation still showed no significant difference between the two periods (insulin: 1.97 +/- 1.16, SU: 2.33 +/- 1.57 ng/ml).(ABSTRACT TRUNCATED AT 250 WORDS)