Picture from Loer CM et al. (2015) Genetics "Cuticle integrity and biogenic amine synthesis in Caenorhabditis elegans ...."

(B) ptps-1-GFP reporters. Left: head showing NSM expression plus other neurons and nonneuronal cells, including anterior intestine (dorsal-ventral view, maximum intensity projection) (MIP); strain OH11619, described by Zhang et al. (2014). Right: vulval region showing expression in neurons VC4, 5 (weak) and HSN, plus lateral seam cells (lateral view, MIP); strain CZ18321.

Picture from Ho J et al. (2018) MicroPubl Biol "Characterizing Dyneins Role in P-cell Nuclear Migration using an Auxin-Induced ...."

Figure 1. Auxin-induced degradation of dynein causes a P-cell nuclear migration defect. Average number of GABA neurons in L4 animals that express the TIR1 gene (an auxin-induced E3 ubiquitin ligase; Zhang et al., 2015) specifically in P cells. Animals either expressed the DHC-1 degron tag (UD551) or did not express the DHC-1 degron tag (UD550). These animals (UD551 and UD550) were either exposed to auxin (+) or not (-) as L1 animals. Statistical significance calculated by t-test with p-value<0.0001. The mean is marked with error bars denoting 95% CI.

Picture from Clark, James F et al. MicroPubl Biol

C. elegans contain three 9 fatty acid desaturases (FADs), fat-5, fat-6, and fat-7, responsible for the initial desaturation step in the synthesis of complex polyunsaturated fatty acids (PUFAs). Production of PUFAs is vital for survival, as simultaneous loss-of-function in all three genes results in synthetic lethality (Brock et al. 2006). Animals containing mutations in any one of the three 9 FADs, fat-5, fat-6, and fat-7, display an increase in body length. fat-5 mutants display a significant increase from 48 hours onward, while fat-6 and fat-7 display a significant increase from 72 hours onward. Since fat-6 and fat-7 mutants have reduced lipid accumulation (Horikawa et al. 2008; Zhang et al. 2013; Clark et al. 2018), these findings on body size suggest that the two phenotypes are not correlated. Consistent with this conclusion, dbl-1 mutants display both decreased body size and lipid accumulation while lon-2 and dbl-1(OE) mutants display increased body size and decreased lipid accumulation at the L4 stage (Clark et al. 2018). Error bars denote SEM, Statistical significance determined via Student's T-test, n.s. not significant, * p<0.05, ** p<.0.01,*** p<0.001. n>25 per strain, repeated in duplicate. Animals were synchronized via an egg lay followed by a timed hatch. Animals were collected and imaged at 25X using a QImaging Retiga EXi camera with QCapture software at each timepoint. Body length was measured by tracing the midline of the animals in ImageJ.

Picture from Soto, Jose et al. (2020) MicroPubl Biol

Principle of the FRET-based ATP sensor, after Zhang et al., 2018. When not bound to ATP, the mApple and Clover fluorochromes are physically far apart. Clover is a form of green fluorescent protein that is excited by blue light and emits green light. mApple is a form of red fluorescent protein that is excited by green light and emits red light. The blue excitation light used for Clover produces only minimal excitation of mApple. However, when ATP is bound, the two fluorescent portions of the sensor are brought close together, permitting emission from Clover to directly excite mApple by FRET. Hence, the amount of red emission from the sensor protein, due to blue excitation, is an indirect measurement of the fraction of protein that is bound to ATP. As a control, excitation/emission of the Clover or mApple portions can detect the sensor protein independent of its interaction with ATP. In our hands, detection of mApple produced a much higher signal-to-noise ratio because of lower red autofluorescence in the gut. B. Image acquisition and analysis. The anterior gut was imaged twice with a 63x objective. The first image used direct excitation and emission of mApple using a TRITC filter set to assess where the sensor protein was present. The second image was obtained using a FRET filter set. This allows excitation of Clover, which produces green emission that excites mApple which is detected as red emission. Because the FRET emissions are of a much lower intensity, the image shown here was enhanced for contrast. Using a Python script, the TRITC image was used to identify 100x100 pixel blocks that contain the sensor protein, and hence define a region of interest (ROI). Pixel values were obtained in the red channel for the TRITC and FRET images across the ROI to generate an average red pixel value ratio. Abbreviations: ph, pharynx; int1, first intestinal ring. C. Histograms showing the data for control and test cases. Error bars denote SEM. (a.u. = arbitrary units)

Picture from Grubbs, Jeremy J. et al. MicroPubl Biol

Figure 1: Regulation of sleep by KIN-29 is not developmental. A) Schematic of the CRISPR/Cas9 strain construction with the DEGRON and GFP-tags inserted at the end of the kin-29 open reading frame. For panels B, D, E, G and H, the average movement quiescence in minutes +/- SEM are displayed following exposure to 1500 J/m2 UVC irradiation. (+) indicates that 2 mM auxin was present during development and analysis. B) UVC-SIS is similar to wild type (WT) in auxin-exposed tir1::mRuby and kin-29::degron::GFP animals, and in tir1::mRuby; kin-29::degron::GFP (KIN-29 AID) transgenic animals in the absence of auxin. C) An image of a KIN-29 AID L4 stage animal. GFP is expressed in head neurons (white arrow) and in the intestine (white arrow head) consistent with previous reports (Lanjuin and Sengupta 2002). tir1::mRuby is expressed ubiquitously, as reported (Zhang et al. 2015). D) UVC-SIS is impaired in KIN-29 AID animals exposed to auxin from the time of hatching. E) The UVC-SIS impairment produced by continuous auxin-induced degradation of KIN-29 is similar to that of kin-29(oy38) null mutants. F) Images of adult KIN-29 AID animals never exposed to auxin (top left), exposed to auxin since hatching (top right), exposed to auxin for 4 hours as adults (bottom left), and removed from auxin for 12 hours starting at the L4 stage after exposure from hatching (bottom right). White arrows point to head neurons (green), which are visible only in the absence of auxin. Images were captured with identical camera and microscope settings. G) KIN-29 AID animals show a similar defect in UVC-SIS whether exposed to auxin at hatch or from the adult stage for four hours. H) UVC-SIS in KIN-29 AID animals on auxin from hatching through adulthood (+) or until the L4 stage, and examined 12 hours later (-12 hours). Reduction of auxin-induced degradation of KIN-29 during adulthood partially restores SIS behavior. ns = not significant, *P<0.05, **P<0.01, ***P<0.001, ****P< 0.0001, two-way ANOVA or mixed-model analysis with Tukey correction for multiple comparisons. Comparisons are made against wild-type (WT) unless otherwise indicated.

Picture from Loer CM et al. (2015) Genetics "Cuticle integrity and biogenic amine synthesis in Caenorhabditis elegans ...."

Figure 8. Biopterin synthetic genes are expressed in biogenic amine neurons and in the epidermis. (A) Expression of cat-4 GFP reporters in larval stage 2 (L2) worms, construct with 2.7 kb upstream sequence (strain CZ9719). Top: Superficial focal plane showing epidermal expression, especially in the hyp7 syncytium. Seam cells have undergone doubling division and can be seen along the lateral side as darker regions among the brightly staining hyp7 cell. Dendritic endings of CEP neurons can be seen at the tip of the 'nose.' Middle: Medial focal plane showing anal cells, strongly expressing tail epidermal cells and posterior intestinal cells expressing GFP. Bottom: Medial focal plane showing epidermal expression in the body and head. In these transgenics, expression in the head is seen in hyp6, but not the more anterior hyp5. NSM and CEP neuron somas are seen in the head, plus some neuronal processes (especially CEP processes). A few other neuronal somas stain less brightly. The anterior intestine also shows GFP expression, as do some rectal epithelial cells (likely B and Y cells). (B) ptps-1-GFP reporters. Left: head showing NSM expression plus other neurons and nonneuronal cells, including anterior intestine (dorsal-ventral view, maximum intensity projection) (MIP); strain OH11619, described by Zhang et al. (2014). Right: vulval region showing expression in neurons VC4, 5 (weak) and HSN, plus lateral seam cells (lateral view, MIP); strain CZ18321. (C) pcbd-1 reporter in larva showing expression in epidermal syncytium hyp7, tail and head epidermis, but not in seam cells; strain CZ19212. Out of focus: pharyngeal muscle and rectal epithelium expression. Inset: enlargement of midbody region. (D) qdpr-1 reporters. Top images: single optical sections from larva showing broad epidermal expression, but little expression in neurons; strain CZ19213. Insets: enlargements of midbody regions. Bottom image: Head showing expression in identified 5HT and DA neurons: NSMs, ADFs, CEPs, and other cells, with reduced epidermal expression; strain CZ19215 (lateral view, MIP). (E) gfrp-1 reporter showing expression in epidermal syncytium hyp7 (top: superficial focal plane) and other epidermal cells (bottom: central focal plane). (A-E) Anterior is to the left in all worms. (A, C, and E) Standard epifluorescence. (B and D) Laser scanning confocal imaging; (B and D, bottom) images are maximal intensity projection of Z-stack; and (D, top and middle) single confocal image planes.