The transcription factor NF-κB plays an important role in the immune system, apoptosis and inflammation. Dorsal, a homolog of NF-κB, patterns the dorsal-ventral axis in the blastoderm embryo. During this stage, Dorsal is sequestered outside the nucleus by the IκB homolog Cactus. Toll signaling on the ventral side breaks the Dorsal/Cactus complex, allowing Dorsal to enter the nucleus to regulate target genes. Fluorescent data show that Dorsal accumulates on the ventral side of the syncytial blastoderm. Here, we use modeling and experimental studies to show that this accumulation is caused by facilitated diffusion, or shuttling, of the Dorsal/Cactus complex. We also show that active Toll receptors are limiting in wild-type embryos, which is a key factor in explaining global Dorsal gradient formation. Our results suggest that shuttling is necessary for viability of embryos from mothers with compromised levels. Therefore, Cactus not only has the primary role of regulating Dorsal nuclear import, but also has a secondary role in shuttling. Given that this mechanism has been found in other, independent, systems, we suggest that it might be more prevalent than previously thought.

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The pheromone response pathway of the yeast S. cerevisiae is a well-established model for the study of G proteins and mitogen-activated protein kinase (MAPK) cascades. Our longstanding ability to combine sophisticated genetic approaches with established functional assays has provided a thorough understanding of signaling mechanisms and regulation. In this report we compare new and established methods used to quantify pheromone-dependent MAPK phosphorylation, transcriptional induction, mating morphogenesis, and gradient tracking. These include both single-cell and population-based assays of activity. We describe several technical advances, provide example data for benchmark mutants, highlight important differences between newer and established methodologies, and compare the advantages and disadvantages of each as applied to the yeast model. Quantitative measurements of pathway activity have been used to develop mathematical models and reveal new regulatory mechanisms in yeast. It is our expectation that experimental and computational approaches developed in yeast may eventually be adapted to human systems biology and pharmacology.

Most aggressive lymphomas are treated with combination chemotherapy, commonly as multiple cycles of concurrent drug administration. Concurrent administration is in theory optimal when combination therapies have synergistic (more than additive) drug interactions. We investigated pharmacodynamic interactions in the standard 4 drug CHOP regimen in Peripheral T Cell Lymphoma (PTCL) cell lines, and found that CHOP consistently exhibits antagonism and not synergy. We tested whether staggered treatment schedules could improve tumor cell kill by avoiding antagonism, using month long in vitro models of concurrent or staggered treatments. Surprisingly, we observed that tumor cell kill is maximized by concurrent drug administration despite antagonistic drug drug interactions. We propose that an ultrasensitive dose response, as described in radiology by the linear quadratic (LQ) model, can reconcile these seemingly contradictory experimental observations. The LQ model describes the relationship between cell survival and dose, and in radiology has identified scenarios favoring hypofractionated radiation: the administration of fewer large doses rather than multiple smaller doses. Specifically, hypofractionated treatment can be favored when cells require an accumulation of DNA damage, rather than a single hit, in order to die. By adapting the LQ model to combination chemotherapy and accounting for tumor heterogeneity, we find that tumor cell kill is maximized by concurrent administration of multiple drugs, even when chemotherapies have antagonistic interactions. Thus, our study identifies a new mechanism by which combination chemotherapy can be clinically beneficial that is not reliant on positive drug drug interactions.