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Open Access Publications from the University of California

Scholarly Works (8 results)

  • Thesis
  • Peer Reviewed
UCLA Electronic Theses and Dissertations (2018)

How multicellular organisms modulate transcription in response to internal and external cues is a fundamental question in plant and animal development and growth. In Arabidopsis, TOPLESS(TPL), a Groucho/Tup1 like co-repressor, is involved in the transcriptional response to multiple plant hormones and plays a crucial role in a range of major developmental processes, such as embryonic apical-basal patterning and post-embryonic floral patterning. This dissertation describes the advances made toward the elucidation of the molecular mechanisms underlying transcriptional regulation by TPL.

As a co-repressor, TPL lacks intrinsic DNA binding activity and has been shown to be recruited by transcription factors with repressive motifs. To date, a range of transcription factors have been reported to interact with TPL, yet the functionality of most of these interactions is not known. We found that the members of the Class III Homedomain-Leucine Zipper (HD- ZIPIII) transcription factor family co-localize with TPL to form complexes at the promoters of thousands of genes in the genome. HD-ZIPIIIs both antagonistically and cooperatively regulate entire pathways at multiple steps in a cell or tissue specific manner. The five members in the family form homo- and hetero-dimers and act both as activators and repressors. The association with TPL confers the repressive function of HD-ZIPIIIs.

Just like its counterparts in animals and fungi, TPL also associates with histone deacetylases(HDACs) to repress transcription. We characterized the binding profiles of TPL and HDA19, a class I histone deacetylase that interacts with TPL, and found they tend to bind to the promoters of dynamically expressed genes. Almost all HDA19 peaks have an overlapping TPL peak, while about half of the TPL peaks are in common with HDA19, suggesting HDA19 is involved only at a subset of TPL targets. Specifically, we found the three members of the evolutionarily conserved NAM/CUC (for NO APICAL MERISTEM and CUP-SHAPED COTYLEDON) transcription factors, CUC1, CUC2 and CUC3, are regulated by TPL and HDA19 to control boundary specification during floral development.

Finally, we showed that TPL interacts with lysine 27-trimethylated histone H3(H3K27me3) and binds to a significant number of genomic regions with this modification, suggesting a novel layer of regulation which has not been reported in animals. A decrease in TPL binding was observed in plants that have a mutation in the Polycomb Repressive Complexes 2 (PRC2), which mediates the deposition of H3K27me3. TPL itself affects H3K27me3 levels and interacts with a component of the PRC1 complex, which recognizes H3K27me3 and further contributes to chromatin compaction. Genes potentially regulated by both TPL and PRCs are involved in multiple developmental processes and stress responses. These data suggest that upon the recruitment of TPL, binding affinity is enhanced by H3K27me3 marked nucleosomes and TPL may further contributes to the repressive transcriptional state via the association with Polycomb group (PcG) proteins.

Overall, we propose a model of transcriptional regulation by TPL that consists of three steps: TPL first is recruited by specific transcription factors; the association of TPL at some target genes is enhanced by the presence of H3K27me3; and TPL forms co-repressor complexes with HDA19 and possibly PcGs to repress gene expression.

  • 3 supplemental files
Cover page: On Transcriptional Regulation by TOPLESS in Arabidopsis thaliana
  • Article
  • Peer Reviewed
UCLA Previously Published Works (2013)
DNA methylation is an epigenetic mark that is associated with transcriptional repression of transposable elements and protein-coding genes. Conversely, transcriptionally active regulatory regions are strongly correlated with histone 3 lysine 4 di- and trimethylation (H3K4m2/m3). We previously showed that Arabidopsis thaliana plants with mutations in the H3K4m2/m3 demethylase JUMONJI 14 (JMJ14) exhibit a mild reduction in RNA-directed DNA methylation (RdDM) that is associated with an increase in H3K4m2/m3 levels. To determine whether this incomplete RdDM reduction was the result of redundancy with other demethylases, we examined the genetic interaction of JMJ14 with another class of H3K4 demethylases: lysine-specific demethylase 1-like 1 and lysine-specific demethylase 1-like 2 (LDL1 and LDL2). Genome-wide DNA methylation analyses reveal that both families cooperate to maintain RdDM patterns. ChIP-seq experiments show that regions that exhibit an observable DNA methylation decrease are co-incidental with increases in H3K4m2/m3. Interestingly, the impact on DNA methylation was stronger at DNA-methylated regions adjacent to H3K4m2/m3-marked protein-coding genes, suggesting that the activity of H3K4 demethylases may be particularly crucial to prevent spreading of active epigenetic marks. Finally, RNA sequencing analyses indicate that at RdDM targets, the increase of H3K4m2/m3 is not generally associated with transcriptional de-repression. This suggests that the histone mark itself--not transcription--impacts the extent of RdDM.
    Cover page: Interplay between active chromatin marks and RNA-directed DNA methylation in Arabidopsis thaliana.
    • Article
    • Peer Reviewed
    UCLA Previously Published Works (2013)
    DNA methylation is an epigenetic mark that is associated with transcriptional repression of transposable elements and protein-coding genes. Conversely, transcriptionally active regulatory regions are strongly correlated with histone 3 lysine 4 di- and trimethylation (H3K4m2/m3). We previously showed that Arabidopsis thaliana plants with mutations in the H3K4m2/m3 demethylase JUMONJI 14 (JMJ14) exhibit a mild reduction in RNA-directed DNA methylation (RdDM) that is associated with an increase in H3K4m2/m3 levels. To determine whether this incomplete RdDM reduction was the result of redundancy with other demethylases, we examined the genetic interaction of JMJ14 with another class of H3K4 demethylases: lysine-specific demethylase 1-like 1 and lysine-specific demethylase 1-like 2 (LDL1 and LDL2). Genome-wide DNA methylation analyses reveal that both families cooperate to maintain RdDM patterns. ChIP-seq experiments show that regions that exhibit an observable DNA methylation decrease are co-incidental with increases in H3K4m2/m3. Interestingly, the impact on DNA methylation was stronger at DNA-methylated regions adjacent to H3K4m2/m3-marked protein-coding genes, suggesting that the activity of H3K4 demethylases may be particularly crucial to prevent spreading of active epigenetic marks. Finally, RNA sequencing analyses indicate that at RdDM targets, the increase of H3K4m2/m3 is not generally associated with transcriptional de-repression. This suggests that the histone mark itself--not transcription--impacts the extent of RdDM.
      • Article
      • Peer Reviewed
      UC Irvine Previously Published Works (2020)

      Background

      The development of carotid-cavernous fistulas (CCFs) during surgical recanalization of chronic internal carotid artery occlusion (ICAO) may be secondary to severe ICA dissection rather than a focal tear of the cavernous ICA seen in typical traumatic CCFs. The purpose of this study is to investigate the causal relationship between the CCFs and severe ICA dissections and to characterize technical outcomes after treatment with stenting.

      Methods

      Five patients underwent treatment with self-expanding stents due to intraprocedural CCF and ICA dissection following surgical removal of ICAO plaque. The stents were telescopically placed via true channel of the dissection. Safety of the procedure was evaluated with 30-day stroke and death rate. Procedural success was determined by the efficacy of CCF obliteration and ICAO recanalization with angiography.

      Results

      All CCFs were associated with spiral and long segmental dissection from the cervical to cavernous ICA. After stenting, successful dissection reconstruction with TICI 3 was achieved in all patients, with complete (n = 4) or partial CCF (n = 1) obliteration. No patient had CCF syndrome, stroke, or death during follow-up of 6 to 37 months; but one patient had pulsatile tinnitus, which resolved 1 year later. Angiography at 6 to 24 months demonstrated CCF obliteration in all 5 patients and durable ICA patency in 4 patients.

      Conclusions

      Intraprocedural CCFs with spiral and cervical-to-cavernous ICA dissection during ICAO surgery are dissection-related because of successful obliteration after stenting for dissection reconstruction. Self-expanding stenting through true channel of the dissection, serving as implanting stent-autograft, may be an optimal therapy for the atypical CCF complication from ICAO surgery.
        Cover page: Dissection-related carotid-cavernous fistula (CCF) following surgical revascularization of chronic internal carotid artery occlusion: a new subtype of CCF and proposed management
        • Article
        • Peer Reviewed
        UCLA Previously Published Works (2016)
        Eukaryotic genomes are regulated by epigenetic marks that act to modulate transcriptional control as well as to regulate DNA replication and repair. In Arabidopsis thaliana, mutation of the ATXR5 and ATXR6 histone methyltransferases causes reduction in histone H3 lysine 27 monomethylation, transcriptional upregulation of transposons, and a genome instability defect in which there is an accumulation of excess DNA corresponding to pericentromeric heterochromatin. We designed a forward genetic screen to identify suppressors of the atxr5/6 phenotype that uncovered loss-of-function mutations in two components of the TREX-2 complex (AtTHP1, AtSAC3B), a SUMO-interacting E3 ubiquitin ligase (AtSTUbL2) and a methyl-binding domain protein (AtMBD9). Additionally, using a reverse genetic approach, we show that a mutation in a plant homolog of the tumor suppressor gene BRCA1 enhances the atxr5/6 phenotype. Through characterization of these mutations, our results suggest models for the production atxr5 atxr6-induced extra DNA involving conflicts between the replicative and transcriptional processes in the cell, and suggest that the atxr5 atxr6 transcriptional defects may be the cause of the genome instability defects in the mutants. These findings highlight the critical intersection of transcriptional silencing and DNA replication in the maintenance of genome stability of heterochromatin.
          Cover page: Identification of Multiple Proteins Coupling Transcriptional Gene Silencing to Genome Stability in Arabidopsis thaliana
          • Article
          • Peer Reviewed
          UCLA Previously Published Works (2015)
          JmjC domain containing protein 14 (JMJ14) is an H3K4-specific histone demethylase that plays important roles in RNA-mediated gene silencing and flowering time regulation in Arabidopsis. However, how JMJ14 is recruited to its target genes remains unclear. Here, we show that the C-terminal FYRN and FYRC domains of JMJ14 are required for RNA silencing and flowering time regulation. Chromatin binding of JMJ14 is lost upon deletion of its FYRN and FYRC domains, and H3K4me3 is increased. FYRN and FYRC domains interact with a pair of NAC domain containing transcription factors, NAC050 and NAC052. Genome-wide ChIP analysis revealed that JMJ14 and NAC050/052 share a set of common target genes with CTTGNNNNNCAAG consensus sequences. Mutations in either NAC052 or NAC050 impair RNA-mediated gene silencing. Together, our findings demonstrate an important role of FYRN and FYRC domains in targeting JMJ14 through direct interaction with NAC050/052 proteins, which reveals a novel mechanism of histone demethylase recruitment.
            • Article
            • Peer Reviewed
            UC San Diego Previously Published Works (2014)
            Recent interest has focused on the importance of the nucleus and associated nucleoskeleton in regulating changes in cardiac gene expression in response to biomechanical load. Mutations in genes encoding proteins of the inner nuclear membrane and nucleoskeleton, which cause cardiomyopathy, also disrupt expression of a biomechanically responsive gene program. Furthermore, mutations in the outer nuclear membrane protein Nesprin 1 and 2 have been implicated in cardiomyopathy. Here, we identify for the first time a role for the outer nuclear membrane proteins, Nesprin 1 and Nesprin 2, in regulating gene expression in response to biomechanical load. Ablation of both Nesprin 1 and 2 in cardiomyocytes, but neither alone, resulted in early onset cardiomyopathy. Mutant cardiomyocytes exhibited altered nuclear positioning, shape, and chromatin positioning. Loss of Nesprin 1 or 2, or both, led to impairment of gene expression changes in response to biomechanical stimuli. These data suggest a model whereby biomechanical signals are communicated from proteins of the outer nuclear membrane, to the inner nuclear membrane and nucleoskeleton, to result in changes in gene expression required for adaptation of the cardiomyocyte to changes in biomechanical load, and give insights into etiologies underlying cardiomyopathy consequent to mutations in Nesprin 1 and 2.
              Cover page: Targeted Ablation of Nesprin 1 and Nesprin 2 from Murine Myocardium Results in Cardiomyopathy, Altered Nuclear Morphology and Inhibition of the Biomechanical Gene Response
              • Article
              • Peer Reviewed
              UC Irvine Previously Published Works (2019)

              Background

              An in situ recanalization procedure of endovascular therapy (ET) or carotid endarterectomy (CEA) has been attempted in patients with symptomatic chronic internal carotid artery occlusions (ICAOs), though the recanalization rates of both are low.

              Objective

              To investigate the outcomes of Multimodality In situ Recanalization for ICAOs in a Hybrid Operating Room (MIRHOR) at the same session.

              Methods

              Symptomatic chronic ICAOs were classified into type A or B (short occlusion with or without a tapered residual root [TRR]), and C or D (long occlusion with or without TRR), and managed in a hybrid operating room with ET, CEA, or both, as needed. Primary efficacy outcome was technical success of recanalization with Thrombolysis in Myocardial Infarction 3. Secondary efficacy outcome was any stroke or death within 30 days (primary safety outcome) plus an ipsilateral ischemic stroke after 30 days.

              Results

              Technical success was finally achieved in 35 (83.3%) of 42 consecutively enrolled patients with ICAO, which was significantly higher than 35.7% (15/42, p<0.001) from the initial ET or CEA alone. Furthermore, the success rate was in descending order: 100% (18/18) for type A and B occlusions, 75% (6/8) for type C occlusions, and 69% (11/16) for type D occlusions (p=0.017). Two secondary efficacy outcome events (5.1%) without mortality, including one (2.4%) primary safety outcome, were observed during a mean follow-up of 10.5 months.

              Conclusion

              The MIRHOR for symptomatic chronic ICAOs at the same session significantly improves technical success, with low periprocedural complications and favorable clinical outcomes. The ICAO classification appears valuable in predicting technical success.
                Cover page: Outcomes of Multimodality In situ Recanalization in Hybrid Operating Room (MIRHOR) for symptomatic chronic internal carotid artery occlusions
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