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Epistatic interactions between Tgfb1 and genetic loci, Tgfbm2 and Tgfbm3, determine susceptibility to an asthmatic stimulus

UC San Francisco Previously Published Works (2012)

TGFβ activation and signaling have been extensively studied in experimental models of allergen-induced asthma as potential therapeutic targets during chronic or acute phases of the disease. Outcomes of experimental manipulation of TGFβ activity have been variable, in part due to use of different model systems. Using an ovalbumin (OVA)-induced mouse model of asthma, we here show that innate variation within TGFβ1 genetic modifier loci, Tgfbm2 and Tgfbm3, alters disease susceptibility. Specifically, Tgfbm2(129) and Tgfbm3(C57) synergize to reverse accentuated airway hyperresponsiveness (AHR) caused by low TGFβ1 levels in Tgfb1(+/-) mice of the NIH/OlaHsd strain. Moreover, epistatic interaction between Tgfbm2(129) and Tgfbm3(C57) uncouples the inflammatory response to ovalbumin from those of airway remodeling and airway hyperresponsiveness, illustrating independent genetic control of these responses. We conclude that differential inheritance of genetic variants of Tgfbm genes alters biological responses to reduced TGFβ1 signaling in an experimental asthma model. TGFβ antagonists for treatment of lung diseases might therefore give diverse outcomes, dependent on genetic variation.

Cover page: Epistatic interactions between Tgfb1 and genetic loci, Tgfbm2 and Tgfbm3, determine susceptibility to an asthmatic stimulus

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General quantitative relations linking cell growth and the cell cycle in Escherichia coli

UC San Diego Previously Published Works (2020)

Growth laws emerging from studies of cell populations provide essential constraints on the global mechanisms that coordinate cell growth^1-3. The foundation of bacterial cell cycle studies relies on two interconnected dogmas that were proposed more than 50 years ago-the Schaechter-Maaloe-Kjeldgaard growth law that relates cell mass to growth rate¹ and Donachie's hypothesis of a growth-rate-independent initiation mass⁴. These dogmas spurred many efforts to understand their molecular bases and physiological consequences^5-14. Although they are generally accepted in the fast-growth regime, that is, for doubling times below 1 h, extension of these dogmas to the slow-growth regime has not been consistently achieved. Here, through a quantitative physiological study of Escherichia coli cell cycles over an extensive range of growth rates, we report that neither dogma holds in either the slow- or fast-growth regime. In their stead, linear relations between the cell mass and the rate of chromosome replication-segregation were found across the range of growth rates. These relations led us to propose an integral-threshold model in which the cell cycle is controlled by a licensing process, the rate of which is related in a simple way to chromosomal dynamics. These results provide a quantitative basis for predictive understanding of cell growth-cell cycle relationships.

Cover page: General quantitative relations linking cell growth and the cell cycle in Escherichia coli

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Differential Requirements for eIF4E Dose in Normal Development and Cancer

UC San Francisco Previously Published Works (2015)

eIF4E, the major cap-binding protein, has long been considered limiting for translating the mammalian genome. However, the eIF4E dose requirement at an organismal level remains unexplored. By generating an Eif4e haploinsufficient mouse, we found that a 50% reduction in eIF4E expression, while compatible with normal development and global protein synthesis, significantly impeded cellular transformation. Genome-wide translational profiling uncovered a translational program induced by oncogenic transformation and revealed a critical role for the dose of eIF4E, specifically in translating a network of mRNAs enriched for a unique 5' UTR signature. In particular, we demonstrate that the dose of eIF4E is essential for translating mRNAs that regulate reactive oxygen species, fueling transformation and cancer cell survival in vivo. Our findings indicate eIF4E is maintained at levels in excess for normal development that are hijacked by cancer cells to drive a translational program supporting tumorigenesis.

Cover page: Differential Requirements for eIF4E Dose in Normal Development and Cancer

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Transcription and methylation analyses of preleukemic promyelocytes indicate a dual role for PML/RARA in leukemia initiation

UC San Francisco Previously Published Works (2015)

Acute promyelocytic leukemia is an aggressive malignancy characterized by the accumulation of promyelocytes in the bone marrow. PML/RARA is the primary abnormality implicated in this pathology, but the mechanisms by which this chimeric fusion protein initiates disease are incompletely understood. Identifying PML/RARA targets in vivo is critical for comprehending the road to pathogenesis. Utilizing a novel sorting strategy, we isolated highly purified promyelocyte populations from normal and young preleukemic animals, carried out microarray and methylation profiling analyses, and compared the results from the two groups of animals. Surprisingly, in the absence of secondary lesions, PML/RARA had an overall limited impact on both the transcriptome and methylome. Of interest, we did identify down-regulation of secondary and tertiary granule genes as the first step engaging the myeloid maturation block. Although initially not sufficient to arrest terminal granulopoiesis in vivo, such alterations set the stage for the later, complete differentiation block seen in leukemia. Further, gene set enrichment analysis revealed that PML/RARA promyelocytes exhibit a subtle increase in expression of cell cycle genes, and we show that this leads to both increased proliferation of these cells and expansion of the promyelocyte compartment. Importantly, this proliferation signature was absent from the poorly leukemogenic p50/RARA fusion model, implying a critical role for PML in the altered cell-cycle kinetics and ability to initiate leukemia. Thus, our findings challenge the predominant model in the field and we propose that PML/RARA initiates leukemia by subtly shifting cell fate decisions within the promyelocyte compartment.

Article
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Breast tumor copy number aberration phenotypes and genomic instability

UC San Francisco Previously Published Works (2006)

Background: Genomic DNA copy number aberrations are frequent in solid tumors, although the underlying causes of chromosomal instability in tumors remain obscure. Genes likely to have genomic instability phenotypes when mutated ( e. g. those involved in mitosis, replication, repair, and telomeres) are rarely mutated in chromosomally unstable sporadic tumors, even though such mutations are associated with some heritable cancer prone syndromes. Methods: We applied array comparative genomic hybridization (CGH) to the analysis of breast tumors. The variation in the levels of genomic instability amongst tumors prompted us to investigate whether alterations in processes/genes involved in maintenance and/or manipulation of the genome were associated with particular types of genomic instability. Results: We discriminated three breast tumor subtypes based on genomic DNA copy number alterations. The subtypes varied with respect to level of genomic instability. We find that shorter telomeres and altered telomere related gene expression are associated with amplification, implicating telomere attrition as a promoter of this type of aberration in breast cancer. On the other hand, the numbers of chromosomal alterations, particularly low level changes, are associated with altered expression of genes in other functional classes (mitosis, cell cycle, DNA replication and repair). Further, although loss of function instability phenotypes have been demonstrated for many of the genes in model systems, we observed enhanced expression of most genes in tumors, indicating that over expression, rather than deficiency underlies instability. Conclusion: Many of the genes associated with higher frequency of copy number aberrations are direct targets of E2F, supporting the hypothesis that deregulation of the Rb pathway is a major contributor to chromosomal instability in breast tumors. These observations are consistent with failure to find mutations in sporadic tumors in genes that have roles in maintenance or manipulation of the genome.

Cover page: Breast tumor copy number aberration phenotypes and genomic instability

Article
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Gene Expression Profile Identifies Tyrosine Kinase c-Met as a Targetable Mediator of Antiangiogenic Therapy Resistance

UC San Francisco Previously Published Works (2013)

Purpose

To identify mediators of glioblastoma antiangiogenic therapy resistance and target these mediators in xenografts.

Experimental design

We conducted microarray analysis comparing bevacizumab-resistant glioblastomas (BRG) with pretreatment tumors from the same patients. We established novel xenograft models of antiangiogenic therapy resistance to target candidate resistance mediator(s).

Results

BRG microarray analysis revealed upregulation versus pretreatment of receptor tyrosine kinase c-Met, which underwent further investigation because of its prior biologic plausibility as a bevacizumab resistance mediator. BRGs exhibited increased hypoxia versus pretreatment in a manner correlating with their c-Met upregulation, increased c-Met phosphorylation, and increased phosphorylation of c-Met-activated focal adhesion kinase and STAT3. We developed 2 novel xenograft models of antiangiogenic therapy resistance. In the first model, serial bevacizumab treatment of an initially responsive xenograft generated a xenograft with acquired bevacizumab resistance, which exhibited upregulated c-Met expression versus pretreatment. In the second model, a BRG-derived xenograft maintained refractoriness to the MRI tumor vasculature alterations and survival-promoting effects of bevacizumab. Growth of this BRG-derived xenograft was inhibited by a c-Met inhibitor. Transducing these xenograft cells with c-Met short hairpin RNA inhibited their invasion and survival in hypoxia, disrupted their mesenchymal morphology, and converted them from bevacizumab-resistant to bevacizumab-responsive. Engineering bevacizumab-responsive cells to express constitutively active c-Met caused these cells to form bevacizumab-resistant xenografts.

Conclusion

These findings support the role of c-Met in survival in hypoxia and invasion, features associated with antiangiogenic therapy resistance, and growth and therapeutic resistance of xenografts resistant to antiangiogenic therapy. Therapeutically targeting c-Met could prevent or overcome antiangiogenic therapy resistance.