The Transformation of a Corporate Culture in a Mature Organization

Much recent attention has been focused on organizational culture as a useful vehicle for understanding organizations. There have been many studies of the cultures of organizations and some on efforts by leaders to change them or implant new ones. Most of these have dealt with founders, their successors or other early generations of leadership or management. Many of them are anecdotal and most, if not all, have been conducted by researchers external to the organization. The limited time and access typically available to the researchers have generally impeded efforts to penetrate to the core culture or to identify significant subcultures. Moreover, there is a paucity of research efforts which examine the transformation of an existing culture in an already-mature business enterprise. This descriptive research employs the historical case study, ethnographic and phenomenological methodologies to determine how a culture evolved at Great American First Savings Bank and how it was transformed by leaders to facilitate attainment of a shared vision and goals for the organization. A history of the 104-year-old financial institution has been developed to serve as necessary background for understanding the culture which evolved. The study describes the current corporate culture as well as the several subcultures which have developed and the persons, events and circumstances which influenced them. The interaction of the main culture and subcultures was also analyzed. Examined also was the effect of mergers and acquisitions upon the parent culture and the culture of the acquired companies. Attempts to preserve the main culture during periods of dramatic change, dynamic growth, deregulation and transformation to a sales and service culture were analyzed. Data were gathered from records, minutes, artifacts, newspaper accounts, periodicals, meetings, conversations and direct observations extending over a period of five years. Additionally, 23 interviews were conducted with current and former members of the organization at various levels and in various locations throughout the company. Results indicate that the culture evolved into a strong, coherent culture over a period of 85 years and that two leaders effected a major transformation of it in order to better position the company and its stakeholders for the growth and competitive environment they knew lay ahead. The findings further suggest that the concept of culture modification is a powerful tool for effecting transformational change or for leading organizations through periods of turbulent change while preserving the basic values and spirit of the organization

Development of an In Vitro Compartmentalization Screen for High-Throughput Directed Evolution of [FeFe] Hydrogenases

BACKGROUND: [FeFe] hydrogenase enzymes catalyze the formation and dissociation of molecular hydrogen with the help of a complex prosthetic group composed of common elements. The development of energy conversion technologies based on these renewable catalysts has been hindered by their extreme oxygen sensitivity. Attempts to improve the enzymes by directed evolution have failed for want of a screening platform capable of throughputs high enough to adequately sample heavily mutated DNA libraries. In vitro compartmentalization (IVC) is a powerful method capable of screening for multiple-turnover enzymatic activity at very high throughputs. Recent advances have allowed [FeFe] hydrogenases to be expressed and activated in the cell-free protein synthesis reactions on which IVC is based; however, IVC is a demanding technique with which many enzymes have proven incompatible. METHODOLOGY/PRINCIPAL FINDINGS: Here we describe an extremely high-throughput IVC screen for oxygen-tolerant [FeFe] hydrogenases. We demonstrate that the [FeFe] hydrogenase CpI can be expressed and activated within emulsion droplets, and identify a fluorogenic substrate that links activity after oxygen exposure to the generation of a fluorescent signal. We present a screening protocol in which attachment of mutant genes and the proteins they encode to the surfaces of microbeads is followed by three separate emulsion steps for amplification, expression, and evaluation of hydrogenase mutants. We show that beads displaying active hydrogenase can be isolated by fluorescence-activated cell-sorting, and we use the method to enrich such beads from a mock library. CONCLUSIONS/SIGNIFICANCE: [FeFe] hydrogenases are the most complex enzymes to be produced by cell-free protein synthesis, and the most challenging targets to which IVC has yet been applied. The technique described here is an enabling step towards the development of biocatalysts for a biological hydrogen economy

Methods for high-dimensonal analysis of cells dissociated from cyropreserved synovial tissue

Abstract Background Detailed molecular analyses of cells from rheumatoid arthritis (RA) synovium hold promise in identifying cellular phenotypes that drive tissue pathology and joint damage. The Accelerating Medicines Partnership RA/SLE Network aims to deconstruct autoimmune pathology by examining cells within target tissues through multiple high-dimensional assays. Robust standardized protocols need to be developed before cellular phenotypes at a single cell level can be effectively compared across patient samples. Methods Multiple clinical sites collected cryopreserved synovial tissue fragments from arthroplasty and synovial biopsy in a 10% DMSO solution. Mechanical and enzymatic dissociation parameters were optimized for viable cell extraction and surface protein preservation for cell sorting and mass cytometry, as well as for reproducibility in RNA sequencing (RNA-seq). Cryopreserved synovial samples were collectively analyzed at a central processing site by a custom-designed and validated 35-marker mass cytometry panel. In parallel, each sample was flow sorted into fibroblast, T-cell, B-cell, and macrophage suspensions for bulk population RNA-seq and plate-based single-cell CEL-Seq2 RNA-seq. Results Upon dissociation, cryopreserved synovial tissue fragments yielded a high frequency of viable cells, comparable to samples undergoing immediate processing. Optimization of synovial tissue dissociation across six clinical collection sites with ~ 30 arthroplasty and ~ 20 biopsy samples yielded a consensus digestion protocol using 100 μg/ml of Liberase™ TL enzyme preparation. This protocol yielded immune and stromal cell lineages with preserved surface markers and minimized variability across replicate RNA-seq transcriptomes. Mass cytometry analysis of cells from cryopreserved synovium distinguished diverse fibroblast phenotypes, distinct populations of memory B cells and antibody-secreting cells, and multiple CD4+ and CD8+ T-cell activation states. Bulk RNA-seq of sorted cell populations demonstrated robust separation of synovial lymphocytes, fibroblasts, and macrophages. Single-cell RNA-seq produced transcriptomes of over 1000 genes/cell, including transcripts encoding characteristic lineage markers identified. Conclusions We have established a robust protocol to acquire viable cells from cryopreserved synovial tissue with intact transcriptomes and cell surface phenotypes. A centralized pipeline to generate multiple high-dimensional analyses of synovial tissue samples collected across a collaborative network was developed. Integrated analysis of such datasets from large patient cohorts may help define molecular heterogeneity within RA pathology and identify new therapeutic targets and biomarkers