The sequence was determined of 6493 nucleotides encompassing the bet genes of Escherichia coli which encode the osmoregulatory choline-glycine betaine pathway. Four open reading frames were identified: betA encoding choline dehydrogenase, a flavoprotein of 61.9kDa; betB encoding betaine aldehyde dehydrogenase (52.8kDa); betT encoding a proton-motive-force-driven, high-affinity transport system for choline (75.8kDa); and betl, capable of encoding a protein of 21.8kDa, implicated as a repressor involved in choline regulation of the bet genes. Identification of the genes was supported by subcloning, physical mapping of lambda placMu53 insertions, amino acid sequence similarity, or N-terminal amino acid sequencing. The bet genes are tightly spaced, with betT located upstream of, and transcribed divergently to, the tandemly linked betIBA genes.

No abstract

The bet regulon allows Escherichia coli to synthesize the osmoprotectant glycine betaine from choline. It comprises a regulatory gene, betI, and three structural genes: betT (choline porter), betA (choline dehydrogenase), and betB (betaine aldehyde dehydrogenase). The bet genes are regulated by oxygen, choline, and osmotic stress. Primer extension analysis identified two partially overlapping promoters which were responsible for the divergent expression of the betT and betIBA transcripts. The transcripts were initiated 61 bp apart. Regulation of the promoters was investigated by using cat (chloramphenicol acetyltransferase) and lacZ (␤-galactosidase) operon fusions. Mutation of betI on plasmid F2 revealed that BetI is a repressor which regulates both promoters simultaneously in response to the inducer choline. Both promoters remained inducible by osmotic stress in a betI mutant background. On the basis of experiments with hns and hns rpoS mutants, we conclude that osmoregulation of the bet promoters was hns independent. The bet promoters were repressed by ArcA under anaerobic growth conditions. An 89-bp promoter fragment, as well as all larger fragments tested, which included both transcriptional start points, displayed osmotic induction and BetI-dependent choline regulation when linked with a cat reporter gene on plasmid pKK232-8. Flanking DNA, presumably on the betT side of the promoter region, appeared to be needed for ArcA-dependent regulation of both promoters.Hyperosmotically stressed cells of Escherichia coli build up the cytoplasmic osmolarity by accumulation of potassium glutamate and various osmoprotectants (12). The highest osmotolerance is achieved by the accumulation of glycine betaine (hereafter called betaine) (49). Betaine can either be taken up by the ProU and ProP systems (7,8,37) or be synthesized by the Bet system, i.e., the choline-to-betaine pathway (31). Synthesis of betaine requires an external supply of choline or the intermediate metabolite betaine aldehyde. At low external concentrations, choline is mainly taken up by the high-affinity choline porter BetT (K m ϭ 8 M), whereas at higher concentrations choline is also taken up by ProU (K m ϭ 1.5 mM) (30, 50). Oxygen-dependent choline dehydrogenase (BetA) catalyzes both steps in the oxidation of choline to betaine by the way of betaine aldehyde, whereas NAD-dependent betaine aldehyde dehydrogenase (BetB) is specific for the last step (1, 31).Biochemical data have previously revealed that expression of the Bet system is reduced under anaerobic conditions. For aerobic cells, osmotic stress gives a partial induction, but for full expression, the cells also need an external supply of choline (31). Experiments with lacZ fusions showed that the regulation occurs at the level of transcription (16).The DNA sequence of the bet region has revealed that in addition to the structural genes, the bet system encodes a regulatory protein called BetI. Albeit BetI shares some sequence homology with the TetR family of bacterial regulatory proteins (29), Bet...

Adrenal glands of foetal sheep of 40 days gestation to term were incubated with and without ACTH or an increased [K+]. With ACTH, the 40 day foetal adrenal was capable of producing more cortisol and aldosterone per g body weight than was the term adrenal. ACTH was a potent stimulus to aldosterone and cortisol production in foetuses aged 60\p=n-\90days, and this effect declined significantly in the 91\p=n-\120day period. An increased [K+] was stimulatory to aldosterone production only after 120 days gestation. Peripheral blood levels of aldosterone, corticosterone, cortisol, 11-deoxycortisol and 11-deoxycorticosterone were measured in foetuses 60 days to term and the levels of aldosterone and cortisol were significantly lower in 90\p=n-\120day foetuses than in the younger or older ones. Direct adrenal vein cannulation proved all five steroids to be secretory products of the foetal adrenal.The ovine foetal cortex has been shown to play a significant role in the onset of parturition in this species (Liggins 1968) and for this reason its ability to