👤 Darrin D Stuart

The co-occurrence of plasma cell neoplasm (PCN) and lymphoplasmacytic lymphoma (LPL) is rare, and their clonal relationship remains unclear. To evaluate the clinicopathologic characteristics of concomitant LPL/PCN. Retrospectively analyzed clinical and laboratory data of 14 cases. Three patients initially presented with immunoglobulin (Ig) M paraprotein, 1 with IgG paraprotein, and 10 had simultaneous diagnoses of PCN and LPL. In 13 cases, flow cytometry detected both LPL and PCN in marrow biopsies. Furthermore, immunohistochemistry highlighted the 2 neoplastic populations, demonstrating an increased proportion of plasma cells and their expression of cyclin D1, CD56, and/or a non-IgM isotype restriction. All cases exhibited discordant heavy-chain isotypes between LPL and PCN. Thirteen of the 14 cases (92.9%) had concordant light-chain restrictions between the 2 neoplasms, and the remaining case (7.1%) showed discordant light-chain restrictions. Of the 12 patients with follow-up, 5 were treated with myeloma regimens, 2 with LPL regimens, 3 with combined therapy, and 2 with observation alone. Follow-up ranged from 2 to 146 months (median, 12.5 months). One patient died of PCN progression, one died of comorbidity, and 10 patients were alive with or without disease. Survival analysis showed no significant difference from the control. The discordant heavy-chain isotype restrictions between PCN and LPL suggest biclonal B-cell neoplasms, which is supported by PCN's phenotypic distinction, such as the expression of cyclin D1 and/or CD56. However, our series exhibited a tendency toward concordant light-chain restrictions between the 2 neoplasms, raising the possibility that PCN may evolve from LPL through class switching. Show less

Adenylyl cyclase (AC)-stimulated cAMP plays a key role in modulating transport and channel activity along the nephron. However, the role of individual adenylyl cyclase isoforms in such regulation is largely unknown. Since adenylyl cyclase 3 (AC3) is expressed throughout nephron, we investigated its role in the physiologic regulation of renal Na(+) and water transport. Mice were generated with inducible nephron knockout of AC3 (AC3 KO) by breeding mice with loxP-flanked critical exons in the Adcy3 gene with mice expressing Pax8-rtTA and LC-1 transgenes. After doxycycline treatment at 1 month of age, nephron AC3 KO mice had 100% Adcy3 gene recombination in all renal tubule segments, but not in glomeruli. Sodium intake, urinary Na(+) excretion, glomerular filtration rate, and blood pressure were similar between nephron KO mice and the controls during normal, high, and low Na(+) diets. Plasma renin concentration was not different between the two groups during varied Na(+) intake. Moreover, there were no differences in urine volume and urine osmolality between the two genotypes during normal or restricted water intake. In conclusion, these data suggested that AC3 is not involved in the physiological regulation of nephron Na(+) and water handling. Show less

In selecting a method to produce a recombinant protein, a researcher is faced with a bewildering array of choices as to where to start. To facilitate decision-making, we describe a consensus 'what to try first' strategy based on our collective analysis of the expression and purification of over 10,000 different proteins. This review presents methods that could be applied at the outset of any project, a prioritized list of alternate strategies and a list of pitfalls that trip many new investigators. Show less

The Structural Proteomics In Europe (SPINE) programme is aimed at the development and implementation of high-throughput technologies for the efficient structure determination of proteins of biomedical importance, such as those of bacterial and viral pathogens linked to human health. Despite the challenging nature of some of these targets, 175 novel pathogen protein structures (approximately 220 including complexes) have been determined to date. Here the impact of several technologies on the structural determination of proteins from human pathogens is illustrated with selected examples, including the parallel expression of multiple constructs, the use of standardized refolding protocols and optimized crystallization screens. Show less

SPINE (Structural Proteomics In Europe) was established in 2002 as an integrated research project to develop new methods and technologies for high-throughput structural biology. Development areas were broken down into workpackages and this article gives an overview of ongoing activity in the bioinformatics workpackage. Developments cover target selection, target registration, wet and dry laboratory data management and structure annotation as they pertain to high-throughput studies. Some individual projects and developments are discussed in detail, while those that are covered elsewhere in this issue are treated more briefly. In particular, this overview focuses on the infrastructure of the software that allows the experimentalist to move projects through different areas that are crucial to high-throughput studies, leading to the collation of large data sets which are managed and eventually archived and/or deposited. Show less

Transcriptional activation of the budding yeast CLN1 and CLN2 genes during the late G1 phase of the cell cycle has been attributed to a positive feedback loop, wherein the transcription of both genes is stimulated by the accumulation of their protein products. We demonstrate that in cycling cells CLN2 does not play a role in determining the timing of its own transcriptional activation. First, we show that CLN3 alone is sufficient to maximally activate CLN2 transcription. Cells that lack functional CLN1 and CLN2 genes activate the CLN2 promoter with the same kinetics and at the same size as cells in which all three CLN genes are functional. In addition, CLN2 transcription is activated with similar kinetics in cells that have CLN2 as their only functional CLN gene and in CLN-deficient cells. Promoter analysis shows that CLN3-dependent activation of CLN2 transcription is directed primarily through the previously identified UAS1 region although another cis-acting region, UAS2, also can contribute to CLN2 activation under some conditions. The ability to activate transcription of CLN2 is not a unique property of CLN3 because ectopically expressed CLN2 can both activate the endogenous CLN2 promoter and induce Start. We propose that failure of the endogenous CLN2 gene to contribute significantly to activation of its own transcription results from its relative effectiveness at inducing Start, cell cycle progression and, subsequently, inactivation of CLN2 expression. Show less

The budding yeast Saccharomyces cerevisiae CLN1, CLN2, and CLN3 genes encode functionally redundant G1 cyclins required for cell cycle initiation. CLN1 and CLN2 mRNAs accumulate periodically throughout the cell cycle, peaking in late G1. We show that cell cycle-dependent fluctuation in CLN2 mRNA is regulated at the level of transcriptional initiation. Mutational analysis of the CLN2 promoter revealed that the major cell cycle-dependent upstream activating sequence (UAS) resides within a 100-bp fragment. This UAS contains three putative SWI4-dependent cell cycle boxes (SCBs) and two putative MluI cell cycle boxes (MCBs). Mutational inactivation of these elements substantially decreased CLN2 promoter activity but failed to eliminate periodic transcription. Similarly, inactivation of SWI4 decreased CLN2 transcription without affecting its periodicity. We have identified a second UAS in the CLN2 upstream region that can promote cell cycle-dependent transcription with kinetics similar to that of the intact CLN2 promoter. Unlike the major CLN2 UAS, this newly identified UAS promotes transcription in cells arrested in G1 by inactivation of cdc28. This novel UAS is both necessary and sufficient for regulated transcription driven by a CLN2 promoter lacking functional SCBs and MCBs. Although this UAS itself contains no SCBs or MCBs, its activity is dependent upon SWI4 function. The characteristics of this novel UAS suggest that it might have a role in initiating CLN2 expression early in G1 to activate the positive feedback loop that drives maximal Cln accumulation. Show less