The nature of nuclear structures that are required to confer transcriptional regulation by distal enhancers is unknown. We show that long-range enhancer-dependent beta-globin transcription is achieved in vitro upon addition of the DNA architectural protein HMG I/Y to affinity-enriched holo RNA polymerase II complexes. In this system, HMG I/Y represses promoter activity in the absence of an associated enhancer and strongly activates transcription in the presence of a distal enhancer. Importantly, nucleosome formation is neither necessary for long-range enhancer regulation in vitro nor sufficient without the addition of HMG I/Y. Thus, the modulation of DNA structure by HMG I/Y is a critical regulator of long-range enhancer function on both DNA and chromatin-assembled genes. Electron microscopic analysis reveals that HMG I/Y binds cooperatively to preferred DNA sites to generate distinct looped structures in the presence or absence of the beta-globin enhancer. The formation of DNA topologies that enable distal enhancers to strongly regulate gene expression is an intrinsic property of HMG I/Y and naked DNA.

The 3' enhancer of the T cell receptor s.chain (TCR~) gene directs the tissue-and stage-specific expression and V(D)J recombination of this gene locus. Using an in vitro system that reproduces TCRoL enhancer activity efficiently, we show that long-range promoter-enhancer regulation requires a T cell-specific repressor complex and is sensitive to DNA topology. In this system, the enhancer functions to derepress the promoter on supercoiled, but not relaxed, templates. We find that the TCRoL promoter is inactivated by a repressor complex that contains the architectural protein HMG I/Y. In the absence of this repressor complex, expression of the TCR~ gene is completely independent of the 3' enhancer and DNA topology. The interaction of the T cell-restricted protein LEF-1 with the TCR~ enhancer is required for promoter derepression. In this system, the TCR~ enhancer increases the number of active promoters rather than the rate of transcription. Thus, long-range enhancers function in a distinct manner from promoters and provide the regulatory link between repressors, DNA topology, and gene activity.[Key Words: TCR genes; transcription; enhancers; HMG I/Y; derepression; DNA topology] Received December 27, 1996; revised version accepted January 14, 1997.The widespread importance of long-range promoterenhancer interactions in tissue-specific and developmentally regulated gene expression has been documented extensively. Some well-known examples include the immunoglobulin, f~-globin, insulin, B-interferon, ~-fetoprotein, ovalbumin, and T-cell receptor genes, as well as the Drosophila alcohol dehydrogenase Adh, fushi tarazu, and ecdysone-regulated glue genes (for review, see Wasylyk 1988). These studies have yielded a wealth of information about enhancer-dependent transcription, but they have not provided mechanistic data to explain how these control elements actually work. For example, it is currently unknown how a gene must be packaged in the nucleus for natural enhancers to function. In this regard, the requirement for chromatin structure to generate long-range transcriptional regulation has been demonstrated in vitro using reconstituted DNA templates (Laybourn and Kadonaga 1992;Barton and Emerson 1994). It also remains to be determined how promoters communicate with distal enhancers. Several models have been proposed: DNA looping by the association of proteins bound at distal sites, DNA tracking by protein translocation, and long-range interactions affected by DNA topology (for review, see Ptashne 1986; Wang and 1Corresponding author.

The transcriptional activity of pBR322 form V DNA template, a topologically unlinked, highly supercoiled molecule having unusual structures around or within coding regions was studied. Significant transcription was observed in vitro from this template despite high levels of supercoiling. An attenuated transcript, initiated accurately from the P4 promoter of rep gene, was observed which indicated pausing of E. coli RNA polymerase within the gene. This pausing could be removed by relieving the torsional stress implying that a supercoil induced structural alteration within the gene was acting as a transcriptional block. A stabilized unusual structure, most likely a cruciform, was found to be responsible for the elongation block. Absence of initiation from the tetR gene was correlated with the unusual structure present within its promoter region in form V DNA. These in vitro studies show that structural alterations within natural DNA could act as transcriptional blocks both at the level of initiation and elongation.

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Polymorphic forms of the DNA duplex with long stretches of structural monotony are known. Several alternating purine-pyrimidine sequences have been shown to adopt left-handed Z-conformation. We report a DNA sequence d(CGCGCGATCGAT)n exhibiting alternating right-handed B and left-handed Z helical conformation after every half a turn. Further, this unusual conformation with change in handedness after every six base pairs was induced at physiological superhelical density.

Employee stock option Plans (ESOPs) have gathered enormous attention in recent decades and have become the most controversial component of the compensation package. Organizations around the globe have been using ESOPs to compensate their employees at managerial and non-managerial levels. While traditionally the stock options were reserved for top management employees, lately there has been strong growth of broad-based plans primarily to increase firm value. Recent literature examining the effects of broad-based stock options are not limited to executive but available for all employees (Core and Guay, 2001; Oyer and Schaefer, 2005; Hallock and Olson, 2010, etc.). However, the shareholders have become increasingly apprehensive about the size and proliferation of adoption of stock option plans. Accordingly, they have been an issue of debate in both academic research and practice circles. The present paper outlines the theoretical foundations behind the use of ESOPs in the compensation mix and strives to address the controversy of whether or not stock options adoptions result in enhancement in firm value. Though the evidence is mixed on the implications of ESOPs, however, there exists robust support for a positive interrelationship between the adoption of these plans and firm performance for large sized firms.

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Structure at the polypurine-polypyrimidine sequences flanking the HpaII sites (CCGG) in pBR322 form V DNA was probed employing single-hit analysis using HpaII restriction endonuclease. Reduced cleavage efficiency of HpaII sites flanked by polypurine-polypyrimidine sequences suggested that under high torsional stress these sequences adopt unwound structures rendering these sites insensitive to restriction enzyme cleavage. In addition to polypurine-polypyrimidine sequences. HpaII sites flanked by alternating purine-pyrimidine sequence, a potential motif of left handed Z-DNA, were also found to be resistant to HpaII cleavage. Results obtained from various studies implicating structure sensitivity of restriction endonucleases and methylases were compiled and a direct correlation was observed between the occurrence of altered sites in a domain and its G/C content in pBR322 form V DNA.