A Virtual Necropsy: Applications of 3D Scanning for Marine Mammal Pathology and Education

Stranded large whales represent an opportunity to learn about the anatomy and health of these cryptic free-ranging animals. However, where time and access is frequently limited, law enforcement and management priorities often take precedence over research, outreach, and educational uses. On 14 March 2021, a dead female adult humpback whale was reported stranded on an uninhabited island 15 miles west of Sitka, Alaska. The whale was three-dimensionally scanned using light detection and ranging (LiDAR) and photogrammetry before, during, and at multiple time points after a necropsy, including full decomposition 17 days later (NOAA Fisheries permit 18786-01). These scans were organized and displayed on the site Sketchfab with annotations and made publically available as a “4D virtual necropsy” (the fourth dimension is time). After one month, our user survey indicated widespread interest in the platform by both the local community and worldwide by stranding professionals, researchers, and educators. We are unaware of another 3D scan involving a large whale with soft tissue for teaching, research, or public display, despite the ease of 3D scanning with current technologies and the wide-ranging applications

Expression of a Truncated ATHB17 Protein in Maize Increases Ear Weight at Silking

<div><p><i>ATHB17</i> (AT2G01430) is an Arabidopsis gene encoding a member of the α-subclass of the homeodomain leucine zipper class II (HD-Zip II) family of transcription factors. The ATHB17 monomer contains four domains common to all class II HD-Zip proteins: a putative repression domain adjacent to a homeodomain, leucine zipper, and carboxy terminal domain. However, it also possesses a unique N-terminus not present in other members of the family. In this study we demonstrate that the unique 73 amino acid N-terminus is involved in regulation of cellular localization of ATHB17. The ATHB17 protein is shown to function as a transcriptional repressor and an EAR-like motif is identified within the putative repression domain of ATHB17. Transformation of maize with an ATHB17 expression construct leads to the expression of ATHB17Δ113, a truncated protein lacking the first 113 amino acids which encodes a significant portion of the repression domain. Because ATHB17Δ113 lacks the repression domain, the protein cannot directly affect the transcription of its target genes. ATHB17Δ113 can homodimerize, form heterodimers with maize endogenous HD-Zip II proteins, and bind to target DNA sequences; thus, ATHB17Δ113 may interfere with HD-Zip II mediated transcriptional activity via a dominant negative mechanism. We provide evidence that maize HD-Zip II proteins function as transcriptional repressors and that ATHB17Δ113 relieves this HD-Zip II mediated transcriptional repression activity. Expression of ATHB17Δ113 in maize leads to increased ear size at silking and, therefore, may enhance sink potential. We hypothesize that this phenotype could be a result of modulation of endogenous HD-Zip II pathways in maize.</p></div

List of differentially expressed genes in ear inflorescence in common between two <i>ATHB17</i> events.

<p>Table lists the fold change (Fc), raw and fdr p-values for common genes in each event.</p

List of differentially expressed genes in ear in common between two <i>ATHB17</i> events.

<p>Table lists the fold change (Fc), raw and fdr p-values for common genes in each event.</p

Phenology of <i>ATHB17</i> events and control.

<p>Two independent <i>ATHB17</i> events in three hybrids were used in physiological studies conducted in 2011 and 2012 under standard agricultural practices (SAP) for corn production in the Central Corn Belt. The number of days to 50% silking and anthesis were measured and the number of days between anthesis and silking was calculated (ASI) each year for physiological studies conducted under standard agronomic practices conditions. Differences in phenology between <i>ATHB17</i> events and control were determined using an across year combined analysis using a mixed model ANOVA. N denotes the number of data points included per entry in the statistical analysis. Number of event data points were within ±3 of control data points. Results for individual hybrids per year are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0094238#pone.0094238.s004" target="_blank">Table S2</a>.</p

ATHB17Δ113 can bind both Class II and Class I DNA targets and ATHB17Δ113 containing V182A-Q185A-N186A mutation cannot bind Class II DNA target in <i>in vitro</i> assay (measured by Surface Plasmon Resonance (SPR).

<p>Binding affinities and Kinetic constants of ATHB17Δ113 interacting with Class I and Class II type DNA, measured by Biacore 2000, globally fitted. SPR measurements with Biacore 2000 were at 25°C in HBS-EP, 100 ug/ml BSA (10 mM HEPES pH 7.4, 150 mM NaCl, 3 mM EDTA, 0.005% Tween-20,100 ug/ml BSA). Equilibrium dissociation constant K_D = k_off/k_on.</p

ATHB17Δ113 is localized to nucleus.

<p>Fluorescence micrographs of maize leaf protoplasts transformed with (A) GFP alone, (B) AtCBF3 fused to GFP as positive control for nuclear localization or (C) ATHB17Δ113 fused with GFP. Protoplasts from each construct were analyzed by laser scanning microscopy for GFP and chlorophyll fluorescence and acquired images were merged on bright-field image using the Image examiner (See Materials and Methods for details).</p

Dry matter accumulation in <i>ATHB17</i> events and in control at R1 development stage.

<p>Two independent <i>ATHB17</i> events in three hybrids were used in physiological studies conducted in 2011 and 2012 under standard agricultural practices (SAP) for corn production in the Central Corn Belt. Data shown is an across year combined analysis using a mixed model ANOVA of dry matter accumulation data collected at R1. Ear partitioning coefficient was calculated by dividing ear dry weight by total dry weight and analyzed as described above. LSmean for the events and wild type control plants are shown in the table with respective delta, % delta and P-value. N denotes the number of data points included per entry in the statistical analysis. Number of event data points were within ±3 of control data points. Results for individual hybrids per year are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0094238#pone.0094238.s005" target="_blank">Table S3</a>.</p

Maize HD-Zip II proteins function as transcriptional repressors and ATHB17Δ113 relieves the repression caused by endogenous maize HD-Zip II proteins.

<p>(A) Cells were co-transformed with 4 µg/320,000 cells of each reporter (right side label) and an increasing amount of each HD-Zip II family member (left-to-right for each color: 0.008; 0.04; 0.2; 1; 5 µg/320,000 cells). Error bars are 1 SD. (B) Cells were triple-transformed with 4 µg/320,000 cells of each reporter (right side label), 20 ng of each HD-Zip family member and an increasing amount of ATHB17Δ113 (left-to-right for each color: 0.0, 0.3; 0.6; 1.3; 2.5 µg/320,000 cells). For a given reporter, HD-Zip II, and concentration of ATHB17Δ113, the calculation of standard error relies on the number of biological replicates and the estimated error variance derived from a one-way ANOVA.</p

Full- length ATHB17 protein functions as transcriptional repressor and ATHB17Δ113 can relieve repression caused by full-length ATHB17 protein.

<p>Maize mesophyll protoplasts were transformed (A) with 4 µg cells of reporter (Class II::GUS, Class I::GUS or No BS::GUS) and 0–5 µg cells of effector (Full-length ATHB17) or 5 µg of ATHB17Δ113 and <i>Renilla</i> luciferase (B) with 4 µg reporter (Class II::GUS, Class I::GUS or No BS::GUS), 0–5 µg ATHB17Δ113, and 0 (grey bars) or 0.2 µg (blue bars) of ATHB17 full length. DNA amounts are per 320,000 cells. After 18 h, cells were assayed for GUS and luciferase expression. GUS values were divided by luciferase internal control values for each well and normalized to respective GFP samples. Bars are means and error bars represent 1 SD.</p