Reduction in Nuclear Size by DHRS7 in Prostate Cancer Cells and by Estradiol Propionate in DHRS7-Depleted Cells

Increased nuclear size correlates with lower survival rates and higher grades for prostate cancer. The short-chain dehydrogenase/reductase (SDR) family member DHRS7 was suggested as a biomarker for use in prostate cancer grading because it is largely lost in higher-grade tumors. Here, we found that reduction in DHRS7 from the LNCaP prostate cancer cell line with normally high levels of DHRS7 increases nuclear size, potentially explaining the nuclear size increase observed in higher-grade prostate tumors where it is lost. An exogenous expression of DHRS7 in the PC3 prostate cancer cell line with normally low DHRS7 levels correspondingly decreases nuclear size. We separately tested 80 compounds from the Microsource Spectrum library for their ability to restore normal smaller nuclear size to PC3 cells, finding that estradiol propionate had the same effect as the re-expression of DHRS7 in PC3 cells. However, the drug had no effect on LNCaP cells or PC3 cells re-expressing DHRS7. We speculate that separately reported beneficial effects of estrogens in androgen-independent prostate cancer may only occur with the loss of DHRS7/ increased nuclear size, and thus propose DHRS7 levels and nuclear size as potential biomarkers for the likely effectiveness of estrogen-based treatments

Chemical interrogation of nuclear size identifies compounds with cancer cell line specific effects on migration and invasion

[Image: see text] Background: Lower survival rates for many cancer types correlate with changes in nuclear size/scaling in a tumor-type/tissue-specific manner. Hypothesizing that such changes might confer an advantage to tumor cells, we aimed at the identification of commercially available compounds to guide further mechanistic studies. We therefore screened for Food and Drug Administration (FDA)/European Medicines Agency (EMA)-approved compounds that reverse the direction of characteristic tumor nuclear size changes in PC3, HCT116, and H1299 cell lines reflecting, respectively, prostate adenocarcinoma, colonic adenocarcinoma, and small-cell squamous lung cancer. Results: We found distinct, largely nonoverlapping sets of compounds that rectify nuclear size changes for each tumor cell line. Several classes of compounds including, e.g., serotonin uptake inhibitors, cyclo-oxygenase inhibitors, β-adrenergic receptor agonists, and Na(+)/K(+) ATPase inhibitors, displayed coherent nuclear size phenotypes focused on a particular cell line or across cell lines and treatment conditions. Several compounds from classes far afield from current chemotherapy regimens were also identified. Seven nuclear size-rectifying compounds selected for further investigation all inhibited cell migration and/or invasion. Conclusions: Our study provides (a) proof of concept that nuclear size might be a valuable target to reduce cell migration/invasion in cancer treatment and (b) the most thorough collection of tool compounds to date reversing nuclear size changes specific to individual cancer-type cell lines. Although these compounds still need to be tested in primary cancer cells, the cell line-specific nuclear size and migration/invasion responses to particular drug classes suggest that cancer type-specific nuclear size rectifiers may help reduce metastatic spread

One-Size Does Not Fit All—A Networked Approach to Community-Based Monitoring in Large River Basins

Monitoring methods based on Indigenous knowledge have the potential to contribute to our understanding of large watersheds. Research in large, complex, and dynamic ecosystems suggests a participatory approach to monitoring—that builds on the diverse knowledges, practices, and beliefs of local people—can yield more meaningful outcomes than a “one-size-fits-all” approach. Here we share the results of 12 community-based, participatory monitoring projects led by Indigenous governments and organizations in the Mackenzie River Basin (2015–2018). Specifically, we present and compare the indicators and monitoring methods developed by each of these community-based cases to demonstrate the specificity of place, culture, and context. A scalar analysis of these results suggests that the combination of core (common) indicators used across the basin, coupled with others that are meaningful at local level, create a methodological bricolage—a mix of tools, methods, and rules-in-use that are fit together. Our findings, along with those of sister projects in two other major watersheds (Amazon, Mekong), confront assumptions that Indigenous-led community-based monitoring efforts are too local to offer insights about large-scale systems. In summary, a networked approach to community-based monitoring that can simultaneously engage with local- and watershed-level questions of social and ecological change can address gaps in knowledge. Such an approach can create both practices and outcomes that are useful to local peoples as well as to those engaged in basin-wide governance

Dataset Figure_5: Whi5-GFP intensity versus size and time in a synchronous G1 population

Whi5-GFP intensity as a function of time for the different FOV of elutriated cells. This dataset contains: 1. Raw TIF_images_NADH – These are autofluorescence images taken at each time point for the purposes of calculating cell size. Time points were every 10 minutes except at 30 minutes which had to be discarded due to poor focus. 2. Raw TIF_images_WHI5 – These raw image files correspond to images of Whi5-GFP excited at 1000 nm fpr all 11 time points of different FOV obtained at 3 different z positions (1-3) (0, -0.5 um, + 0.5 um). Whi5-GFP intensity values for each nucleus were taken for the z-position that gave the highest intensity for each nucleus. 3. NADHDATA_with_time plot.xlsx is the analysis of the raw images of auto-fluorescence exciting at 750 nm. The only relevant information for the Figure is in Colume C sheet 1. It is the cell area in total pixels. 4. Whi5Data_BestFocusPlanes_All_FOV.xlsx is the analysis of the Whi5-GFP images for Whi5-GFP intensity vs time and size for each time point (which corresponds to a different FOV)

Dataset Figure_4: Nuclear Whi5-GFP intensity versus time from repeated imaging of the same individual cells

Whi5-GFP intensity as a function of time for the same FOVs. Only small daughter cells in the asynchronous population at time point 0 were quantified as a function of time. The folder 'Image Files' contains raw image files for Whi5-GFP for FOV1,3,4,5 & 6 at each of the 6 time points (0, 20, 40, 60, 80, 100 min). The folder 'Excel_analysis_output' contains the output files for all five FOV (1,3,4,5,6) for 1. fovX_t0 or time 100_whi5_nadh_*.xlsx – at time 0 and time 100 minutes. These files correspond to the analysis of the background auto-fluorescence excited at 750 nm used to calculate size. The only relevant information from these analyses is the Cyto size (fL) column in tab 3 of each file 2. fovX_tX_whi5_*.xlsx Analysis of all time points and all FOV for Whi5-GFP intensities

Dataset Figure_6: Cln3 levels pulse prior to Start

Data and Matlab script 1. FLmean_Cln3_GLU: CSV file containing mean GFP concentration from Cln3-P2A-GFP for single daughter cells grown in glucose. Each cell is followed from birth until a few minutes after bud appearance. Data is organized by columns (one column corresponds to one cell). The first column is the vector of measurement time points (in minutes). 2. Vol_Cln3_GLU: CSV file containing cell volume for each of the daughter cells described above. Data is organized by columns (one column corresponds to one cell). The first column is the vector of measurement time points (in minutes). 3. bud_times: Excel file containing the time of bud appearance for each of the daughter cells described above 4. GP_FLtot_Cln3_GLU_example: Matlab script used to perform Gaussian process regression (see description in Methods) on the total GFP fluorescence for each of the daughter cells described above. The script produces single-cell Cln3 abundance data used to generate Fig. 5c of the main text and Extended Data Figure 4H. NOTE: in order to run, the Matlab script requires the installation of the GPML Matlab toolbox (freely available at http://www.gaussianprocess.org/gpml/code/matlab/doc/)

Dataset Figure_2: Determination of nuclear Whi5 concentration by widefield fluorescence microscopy is confounded by non-proportional scaling of the nuclear-to-cell volume

This dataset contain the data used to determine the nuclear to whole-cell volume for the Nup133-yeGFP strain