Straightening Caenorhabditis elegans images

Motivation: Caenorhabditis elegans, a roundworm found in soil, is a widely studied model organism with about 1000 cells in the adult. Producing high-resolution fluorescence images of C.elegans to reveal biological insights is becoming routine, motivating the development of advanced computational tools for analyzing the resulting image stacks. For example, worm bodies usually curve significantly in images. Thus one must ‘straighten’ the worms if they are to be compared under a canonical coordinate system

Automatic reconstruction of 3D neuron structures using a graph-augmented deformable model

Motivation: Digital reconstruction of 3D neuron structures is an important step toward reverse engineering the wiring and functions of a brain. However, despite a number of existing studies, this task is still challenging, especially when a 3D microscopic image has low single-to-noise ratio and discontinued segments of neurite patterns

Quantifying Phenotypic Variation in Isogenic Caenorhabditis elegans Expressing Phsp-16.2::gfp by Clustering 2D Expression Patterns

Isogenic populations of animals still show a surprisingly large amount of phenotypic variation between individuals. Using a GFP reporter that has been shown to predict longevity and resistance to stress in isogenic populations of the nematode Caenorhabditis elegans, we examined residual variation in expression of this GFP reporter. We found that when we separated the populations into brightest 3% and dimmest 3% we also saw variation in relative expression patterns that distinguished the bright and dim worms. Using a novel image processing method which is capable of directly analyzing worm images, we found that bright worms (after normalization to remove variation between bright and dim worms) had expression patterns that correlated with other bright worms but that dim worms fell into two distinct expression patterns. We have analysed a small set of worms with confocal microscopy to validate these findings, and found that the activity loci in these clusters are caused by extremely bright intestine cells. We also found that the vast majority of the fluorescent signal for all worms came from intestinal cells as well, which may indicate that the activity of intestinal cells is responsible for the observed patterns. Phenotypic variation in C. elegans is still not well understood but our proposed novel method to analyze complex expression patterns offers a way to enable a better understanding

Assembling models of embryo development: Image analysis and the construction of digital atlases

Digital atlases of animal development provide a quantitative description of morphogenesis, opening the path toward processes modeling. Prototypic atlases offer a data integration framework where to gather information from cohorts of individuals with phenotypic variability. Relevant information for further theoretical reconstruction includes measurements in time and space for cell behaviors and gene expression. The latter as well as data integration in a prototypic model, rely on image processing strategies. Developing the tools to integrate and analyze biological multidimensional data are highly relevant for assessing chemical toxicity or performing drugs preclinical testing. This article surveys some of the most prominent efforts to assemble these prototypes, categorizes them according to salient criteria and discusses the key questions in the field and the future challenges toward the reconstruction of multiscale dynamics in model organisms

Efficient and robust shape retrieval from deformable templates

Computer Systems, Imagery and Medi

A 3D digital atlas of C. elegans and its application to single-cell analyses,”

We built a digital nuclear atlas of the newly hatched, first larval stage (l1) of the wild-type hermaphrodite of Caenorhabditis elegans at single-cell resolution from confocal image stacks of 15 individual worms. the atlas quantifies the stereotypy of nuclear locations and provides other statistics on the spatial patterns of the 357 nuclei that could be faithfully segmented and annotated out of the 558 present at this developmental stage. We then developed an automated approach to assign cell names to each nucleus in a threedimensional image of an l1 worm. We achieved 86% accuracy in identifying the 357 nuclei automatically. this computational method will allow high-throughput single-cell analyses of the post-embryonic worm, such as gene expression analysis, or ablation or stimulation of cells under computer control in a high-throughput functional screen. Despite the detailed knowledge of the anatomy of the nematode C. elegans 1 as well as its determined cell lineage 2,3 , the mapped connectivity of its nervous system 4,5 and its sequenced genome 6,7 , we still lack a three-dimensional (3D) digital atlas of positions of nuclei in any postembryonic stage. Such an atlas has several potential applications. First, it provides us with previously unavailable quantitative knowledge about the degree of stereotypy of the positions of nuclei and the specific spatial relationships between different cells. Second, the atlas can serve as a standard template; we can compare any 3D image of a wild-type C. elegans to the atlas and extract the identities of individual nuclei using an automated approach. This is essential for high-throughput analysis of cellular information such as gene expression at single-cell resolution. Such an analysis provides much richer information than does analysis of expression data from a DNA microarray experiment 8,9 as DNA microarrays reveal average expression from tissue or from the entire individual but not the expression in an individual cell. Prior to this study, the anatomy of C. elegans has been described qualitatively by images with a text description or two-dimensional sketches 10 . Early efforts using electron microscopy analyses have resulted in detailed views of the anatomy 10 and even a connectivity graph of the nervous system 4,5 , but to date, manual or automated segmentation of the fine structure of such an ultrahigh-resolution image stack has not been demonstrated. Whereas one might contemplate carrying out such a manual segmentation for a single worm, doing so for enough worms to deliver statistical information on the location of nuclei is effectively impractical. Our method for automatically analyzing individual cells in postembryonic worms complements the similar capability developed previously for the embryo results Building a 3d digital atlas We used DAPI (4,6-diamidino-2-phenylindole) to stain the nuclei of all 558 cells. We used a myo-3:GFP transgene to label the nuclei of the 81 body wall muscle cells and 1 depressor muscle cell. These nuclei were fiducial markers, used by our manual and automated approach to annotate cells. We used a gene encoding monomeric Cherry protein (mCherry) driven by a promoter from a gene of interest to reveal expression in a set of target cells. We collected 3D images of C. elegans at the L1 stage using a Leica confocal microscope To build a standard digital atlas, we first computationally straightened the curved worm body in the 3D image into a rod shape 1

Components Required for the Death and Degradation of the Linker Cell in C. Elegans

Programmed cell death (PCD) is an important process in the development of multicellular organisms. Apoptosis, a form of PCD characterized morphologically by chromatin condensation, membrane blebbing, and cytoplasm compaction, and molecularly by the activation of caspase proteases, has been extensively investigated. Studies in C. elegans, Drosophila, mice, and the developing chick have revealed, however, that developmental PCD also occurs through other mechanisms, morphologically and molecularly distinct from apoptosis. One prominent cell death program, linker cell-type death (LCD), is morphologically conserved, and independent of the key genes that drive apoptosis. Instead, LCD functions, in part, through the stressrelated protein HSF-1, and subsequent upregulation of members of the ubiquitin proteasome system. How exactly HSF-1 is post-translationally regulated to either commit to cell survival or cell death is not currently known. Using a protein interaction screen and classical genetic studies, I propose that the homeodomain protein kinase HPK-1 is required to activate the heat shock function of HSF-1, thereby indirectly inhibiting its cell death role. I hypothesize that PQN-41C, a polyglutamine protein necessary for linker cell death, binds HPK-1 to limit its HSF-1 interactions, and pushes the cell towards death. Downstream of cell death, the linker cell must be phagocytosed and degraded by engulfing cells. This process does not rely on canonical apoptotic factors, so I carried out a forward genetic screen to identify genes involved in corpse degradation. I discovered a key protein network involved in linker cell corpse engulfment and degradation and revealed that two small GTPases, RAB-35 and ARF-6, and their regulators ensure timely phagocytosis and phagosome maturation. I also determined that the caspase CED-3 and its upstream regulator CED-4 are required not for cell death, but for proper cell corpse removal. This new role of caspase in cell corpse disposal offers an alternative function for the role of caspases in cell death, and suggests how apoptotic and non-apoptotic forms of cell death may work together to remove cells during animal development

Pattern Recognition in High-Throughput Zebrafish Imaging

High Throughput (HT) methods are high volume experimental approaches that are common in the fields of the life-sciences. The instrumentation for these methods differs per application. We will focus on the HT methods that are concerned with imaging. The aim of this thesis is to find robust methods for object extraction and analysis. We focus on the Computer Science aspects of such analysis, namely pattern recognition. Pattern Recognition can be seen in the context of object recognition and data mining. Both aspects will be described in this thesis. We present a framework for segmenting and recognizing the objects of interest based on Template Matching. This approach was designed for an application in the HT screening of zebrafish embryos. All proposed methods are fully automated. We further elaborate on the segmentation algorithms to apply these in software that can be used in a HT context to derive measurements. Then we apply the software on a real life problem involving zebrafish infected with Mycobacterium marinum.SmartmixComputer Systems, Imagery and Medi