A genomic approach to the identification and characterization of HOXA13 functional binding elements

HOX proteins are important transcriptional regulators in mammalian embryonic development and are dysregulated in human cancers. However, there are few known direct HOX target genes and their mechanisms of regulation are incompletely understood. To isolate and characterize gene segments through which HOX proteins regulate transcription we used cesium chloride centrifugation-based chromatin purification and immunoprecipitation (ChIP). From NIH 3T3-derived HOXA13-FLAG expressing cells, 33% of randomly selected, ChIP clones were reproducibly enriched. Hox-enriched fragments (HEFs) were more AT-rich compared with cloned fragments that failed reproducible ChIP. All HEFs augmented transcription of a heterologous promoter upon coexpression with HOXA13. One HEF was from intron 2 of Enpp2, a gene highly upregulated in these cells and has been implicated in cell motility. Using Enpp2 as a candidate direct target, we identified three additional HEFs upstream of the transcription start site. HOXA13 upregulated transcription from an Enpp2 promoter construct containing these sites, and each site was necessary for full HOXA13-induced expression. Lastly, given that HOX proteins have been demonstrated to interact with histone deacetylases and/or CBP, we explored whether histone acetylation changed at Enpp2 upon HOXA13-induced activation. No change in the general histone acetylation state was observed. Our results support models in which occupation of multiple HOX binding sites is associated with highly activated genes

Genomic Promoter Analysis Predicts Functional Transcription Factor Binding

Background. The computational identification of functional transcription factor binding sites (TFBSs) remains a major challenge of computational biology. Results. We have analyzed the conserved promoter sequences for the complete set of human RefSeq genes using our conserved transcription factor binding site (CONFAC) software. CONFAC identified 16296 human-mouse ortholog gene pairs, and of those pairs, 9107 genes contained conserved TFBS in the 3 kb proximal promoter and first intron. To attempt to predict in vivo occupancy of transcription factor binding sites, we developed a novel marginal effect isolator algorithm that builds upon Bayesian methods for multigroup TFBS filtering and predicted the in vivo occupancy of two transcription factors with an overall accuracy of 84%. Conclusion. Our analyses show that integration of chromatin immunoprecipitation data with conserved TFBS analysis can be used to generate accurate predictions of functional TFBS. They also show that TFBS cooccurrence can be used to predict transcription factor binding to promoters in vivo

HEF enrichment without protein–protein crosslinking and chromatin purification

<p><b>Copyright information:</b></p><p>Taken from "A genomic approach to the identification and characterization of HOXA13 functional binding elements"</p><p>Nucleic Acids Research 2005;33(21):6782-6794.</p><p>Published online 30 Nov 2005</p><p>PMCID:PMC1301594.</p><p>© The Author 2005. Published by Oxford University Press. All rights reserved</p> () ChIP was performed in HOXA13-FLAG and HOX (−) cells using anti-FLAG agarose. Elimination of crosslinking with DMA preceding formaldehyde crosslinking is represented in indicated lanes [DMA(−)]. HEF1 and HEF2 are enriched upon addition of DMA as well as without DMA in the HOXA13-FLAG versus HOX (−) cells. HEF1 demonstrated a visibly higher signal with DMA versus DMA (−) in the HOXA13-FLAG cells (1.9-fold by BioRad Quantity One analysis) while HEF2 recovery was equal between the samples. () ChIP was performed in HOXA13-FLAG and HOX (−) cell lines using anti-FLAG agarose. CsCl purification of chromatin was eliminated in the indicated samples (−). HEF1 and HEF2 are both enriched in the HOXA13-FLAG cells versus the HOX (−) cells both with and without chromatin purification. There was consistently no product in the HOX (−) sample for HEF1; however, there is a product present in the CsCl purified HEF2 sample

Results of an online, globally-available survey to determine modes of cancer treatment education used by patients with cancer.

e18523 Background: Many patients with cancer (PwC) possess a suboptimal understanding of their chemotherapy treatment plan.1 Accordingly, PwC may seek information regarding both the disease and its treatment from other resources. Purpose: A freely-accessible website was curated by oncology pharmacists to provide evidence-based educational material on chemotherapy treatment plans. Objective: To use the website to deploy an online survey assessing other modes of education routinely accessed by PwC. Methods: Between February 2020 and January 2021, PwC completed an anonymous, English language survey via www.chemoexperts.com. Results: A total of 1,036 self-identified patient users began the survey. The majority were from North America (75%). Most respondents (65%) were female. PwC were asked: “If you received education from a health care professional (HCP), what type of education did you receive?” Multiple responses were allowed. Of the 820 patients who responded to this question, 77.4% (n = 635) were given printed material, 52.3% (n = 429) were taught in clinic, 8% (n = 66) were shown videos, 5.7% (n = 47) participated in an educational class outside of clinic, and 4% (n = 33) were given magazines. Roughly 37% were directed to an internet site (n = 304; 92 to chemoexperts.com, 212 to other internet sites) by a HCP. Patients also reported using blogs (12.7%; n = 111/873 respondents), websites other than chemoexperts (85%; n = 742/873), support groups (28.4%; n = 248/873), and YouTube (15.1%; n = 132/873). Discussion: In-person teaching sessions are vital to understanding treatment plans. While the ongoing COVID-19 pandemic may have lowered the percentage of PwC who are offered teaching, some PwC may still not absorb or retain all there is to know during in-person sessions and seek other resources to bolster their understanding of treatment. Although the majority of users reported receiving printed material, this too may be limited in its ability to meet the needs of all individuals. Knowing PwC may seek varying modes of education, HCPs should direct patients to resources they are familiar with. Conclusions: To our knowledge, this is the first global survey to examine modes of education utilized by PwC. Although PwC may receive teaching in person, many still seek a variety of educational resources outside of clinic. Relying solely on printed material will not meet the needs of some PwC. When educating PwC in clinic, HCPs concerned about the validity of outside materials should direct patients to resources they are familiar with and trust. Reference: Almalki H, Absi A, Alghamdi A, Alsalmi M, Khan M. Analysis of Patient-Physician Concordance in the Understanding of Chemotherapy Treatment Plans Among Patients With Cancer. JAMA Netw Open. 2020 Mar 2;3(3):e200341. </jats:p