Help Your Nonprofit Clients Improve Their Accounting for Capital and In-Kind Donations

Many nonprofit organizations, especially smaller ones, lack skilled financial professionals either on staff or on the board. Auditors and other external CPAs are in an excellent position to help these clients present their financial results in ways that will avoid misunderstanding and confusion on the part of financial statement users both inside and outside the client organization. Two areas that have created special problems for nonprofits are accounting for large capital donations and for in-kind donations. We’ll look at the problems first; then show you accounting solutions you can bring to your clients

Glucocorticoid receptor quaternary structure drives chromatin occupancy and transcriptional outcome

Most transcription factors, including nuclear receptors, are widely modeled as binding regulatory elements as monomers, homodimers, or heterodimers. Recent findings in live cells show that the glucocorticoid receptor NR3C1 (also known as GR) forms tetramers on enhancers, owing to an allosteric alteration induced by DNA binding, and suggest that higher oligomerization states are important for the gene regulatory responses of GR. By using a variant (GRtetra) that mimics this allosteric transition, we performed genome-wide studies using a GR knockout cell line with reintroduced wild-type GR or reintroduced GRtetra. GRtetra acts as a super receptor by binding to response elements not accessible to the wild-type receptor and both induces and represses more genes than GRwt. These results argue that DNA binding induces a structural transition to the tetrameric state, forming a transient higher-order structure that drives both the activating and repressive actions of glucocorticoids.Fil: Paakinaho, Ville. National Institutes of Health; Estados Unidos. University Of Eastern Finland; FinlandiaFil: Johnson, Thomas A.. National Institutes of Health; Estados UnidosFil: Presman, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Hager, Gordon L.. National Institutes of Health; Estados Unido

Genome-wide binding potential and regulatory activity of the glucocorticoid receptor’s monomeric and dimeric forms

A widely regarded model for glucocorticoid receptor (GR) action postulates that dimeric binding to DNA regulates unfavorable metabolic pathways while monomeric receptor binding promotes repressive gene responses related to its anti-inflammatory effects. This model has been built upon the characterization of the GRdim mutant, reported to be incapable of DNA binding and dimerization. Although quantitative live-cell imaging data shows GRdim as mostly dimeric, genomic studies based on recovery of enriched half-site response elements suggest monomeric engagement on DNA. Here, we perform genome-wide studies on GRdim and a constitutively monomeric mutant. Our results show that impairing dimerization affects binding even to open chromatin. We also find that GRdim does not exclusively bind half-response elements. Our results do not support a physiological role for monomeric GR and are consistent with a common mode of receptor binding via higher order structures that drives both the activating and repressive actions of glucocorticoids.Fil: Johnson, Thomas A.. National Institutes of Health; Estados UnidosFil: Paakinaho, Ville. University Of Eastern Finland.; FinlandiaFil: Kim, Sohyoung. National Institutes of Health; Estados UnidosFil: Hager, Gordon L.. National Institutes of Health; Estados UnidosFil: Presman, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; Argentin

Meta-analysis of Chromatin Programming by Steroid Receptors

Transcription factors (TFs) must access chromatin to bind to their response elements and regulate gene expression. A widely accepted model proposes that only a special subset of TFs, pioneer factors, can associate with condensed chromatin and initiate chromatin opening. We previously reported that steroid receptors (SRs), not considered pioneer factors, can assist the binding of an archetypal pioneer, the forkhead box protein 1 (FOXA1), at a subset of receptor-activated enhancers. These findings have been challenged recently, with the suggestion that newly acquired data fully support the prevailing pioneer model. Here, we reexamine our results and confirm the original conclusions. We also analyze and discuss a number of available datasets relevant to chromatin penetration by SRs and find a general consensus supporting our original observations. Hence, we propose that chromatin opening at some sites can be initiated by SRs, with a parallel recruitment of factors often treated as having a unique pioneer function.Fil: Paakinaho, Ville. University Of Eastern Finland.; FinlandiaFil: Swinstead, Erin E.. National Cancer Institute; Estados UnidosFil: Presman, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Grøntved, Lars. Technical University of Denmark; DinamarcaFil: Hager, Gordon L.. National Cancer Institute; Estados Unido

State-to-State Rotational Rate Constants for CO+He: Infrared Double Resonance Measurements and Simulation of the Data Using the SAPT Theoretical Potential Energy Surface

An extensive data set of 54 time-resolved pump-probe measurements was used to examine CO+He rotational energy transfer within the CO v=2 rotational manifold. Rotational levels in the range Ji=2-9 were excited and collisional energy transfer of population to the levels Jf=1-10 was monitored. The resulting data set was analyzed by fitting to numerical solutions of the master equation. State-to-state rate constant matrices were generated using fitting law functions and ab initio theoretical calculations that employed the SAPT potential energy surface of Heijmen et al. [J. Chem. Phys. 107, 9921 (1997)]. Fitting laws based on the modified exponential gap (MEG), statistical power exponential gap (SPEG), and energy corrected sudden with exponential power (ECS-EP) models all yielded acceptable simulations of the kinetic data, as did the theoretical rate constants. However, the latter were unique in their ability to reproduce both our kinetic data and the pressure broadening coefficients for CO+He. These results provide an impressive demonstration of the quality of the symmetry adapted perturbation theory (SAPT) potential energy surface

Power-law behaviour of transcription factor dynamics at the single-molecule level implies a continuum affinity model

Single-molecule tracking (SMT) allows the study of transcription factor (TF) dynamics in the nucleus, giving important information regarding the diffusion and binding behavior of these proteins in the nuclear environment. Dwell time distributions obtained by SMT for most TFs appear to follow bi-exponential behavior. This has been ascribed to two discrete populations of TFs-one non-specifically bound to chromatin and another specifically bound to target sites, as implied by decades of biochemical studies. However, emerging studies suggest alternate models for dwell-time distributions, indicating the existence of more than two populations of TFs (multi-exponential distribution), or even the absence of discrete states altogether (power-law distribution). Here, we present an analytical pipeline to evaluate which model best explains SMT data. We find that a broad spectrum of TFs (including glucocorticoid receptor, oestrogen receptor, FOXA1, CTCF) follow a power-law distribution of dwell-times, blurring the temporal line between non-specific and specific binding, suggesting that productive binding may involve longer binding events than previously believed. From these observations, we propose a continuum of affinities model to explain TF dynamics, that is consistent with complex interactions of TFs with multiple nuclear domains as well as binding and searching on the chromatin template.Fil: Garcia, David A.. National Institutes of Health; Estados UnidosFil: Fettweis, Gregory. National Institutes of Health; Estados UnidosFil: Presman, Diego Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Paakinaho, Ville. University Of Eastern Finland.; FinlandiaFil: Jarzynski, Christopher. University of Maryland; Estados UnidosFil: Upadhyaya, Arpita. University of Maryland; Estados UnidosFil: Hager, Gordon L.. National Institutes of Health; Estados Unido

Actin-dependent intranuclear repositioning of an active gene locus in vivo

Although bulk chromatin is thought to have limited mobility within the interphase eukaryotic nucleus, directed long-distance chromosome movements are not unknown. Cajal bodies (CBs) are nuclear suborganelles that nonrandomly associate with small nuclear RNA (snRNA) and histone gene loci in human cells during interphase. However, the mechanism responsible for this association is uncertain. In this study, we present an experimental system to probe the dynamic interplay of CBs with a U2 snRNA target gene locus during transcriptional activation in living cells. Simultaneous four-dimensional tracking of CBs and U2 genes reveals that target loci are recruited toward relatively stably positioned CBs by long-range chromosomal motion. In the presence of a dominant-negative mutant of β-actin, the repositioning of activated U2 genes is markedly inhibited. This supports a model in which nuclear actin is required for these rapid, long-range chromosomal movements