Background

Cryo-electron microscopy (Cryo-EM) and tomography (Cryo-ET) have emerged as important imaging techniques for studying structures of macromolecular complexes. In 3D reconstruction of large macromolecular complexes, many 2D projection images of macromolecular complex particles are usually acquired with low signal-to-noise ratio. Therefore, it is meaningful to select multiple images containing the same structure with identical orientation. The selected images are averaged to produce a higher-quality representation of the underlying structure with improved resolution. Existing approaches of selecting such images have limited accuracy and speed.

Methods

We propose a simulated annealing-based algorithm (SA) to pick the homogeneous image set with best average. Its performance is compared with two baseline methods based on both 2D and 3D datasets. When tested on simulated and experimental 3D Cryo-ET images of Ribosome complex, SA sometimes stopped at a local optimal solution. Restarting is applied to settle this difficulty and significantly improved the performance of SA on 3D datasets.

Results

Experimented on simulated and experimental 2D Cryo-EM images of Ribosome complex datasets respectively with SNR = 10 and SNR = 0.5, our method achieved better accuracy in terms of F-measure, resolution score, and time cost than two baseline methods. Additionally, SA shows its superiority when the proportion of homogeneous images decreases.

Conclusions

SA is introduced for homogeneous image selection to realize higher accuracy with faster processing speed. Experiments on both simulated and real 2D Cryo-EM and 3D Cryo-ET images demonstrated that SA achieved expressively better performance. This approach serves as an important step for improving the resolution of structural recovery of macromolecular complexes captured by Cryo-EM and Cryo-ET.

Cryo-electron Tomography (cryo-ET) generates 3D visualization of cellular organization that allows biologists to analyze cellular structures in a near-native state with nano resolution. Recently, deep learning methods have demonstrated promising performance in classification and segmentation of macromolecule structures captured by cryo-ET, but training individual deep learning models requires large amounts of manually labeled and segmented data from previously observed classes. To perform classification and segmentation in the wild (i.e., with limited training data and with unseen classes), novel deep learning model needs to be developed to classify and segment unseen macromolecules captured by cryo-ET. In this paper, we develop a one-shot learning framework, called cryo-ET one-shot network (COS-Net), for simultaneous classification of macromolecular structure and generation of the voxel-level 3D segmentation, using only one training sample per class. Our experimental results on 22 macromolecule classes demonstrated that our COS-Net could efficiently classify macromolecular structures with small amounts of samples and produce accurate 3D segmentation at the same time.

Salient segmentation is a critical step in biomedical image analysis, aiming to cut out regions that are most interesting to humans. Recently, supervised methods have achieved promising results in biomedical areas, but they depend on annotated training data sets, which requires labor and proficiency in related background knowledge. In contrast, unsupervised learning makes data-driven decisions by obtaining insights directly from the data themselves. In this paper, we propose a completely unsupervised self-aware network based on pre-training and attentional backpropagation for biomedical salient segmentation, named as PUB-SalNet. Firstly, we aggregate a new biomedical data set from several simulated Cellular Electron Cryo-Tomography (CECT) data sets featuring rich salient objects, different SNR settings and various resolutions, which is called SalSeg-CECT. Based on the SalSeg-CECT data set, we then pre-train a model specially designed for biomedical tasks as a backbone module to initialize network parameters. Next, we present a U-SalNet network to learn to selectively attend to salient objects. It includes two types of attention modules to facilitate learning saliency through global contrast and local similarity. Lastly, we jointly refine the salient regions together with feature representations from U-SalNet, with the parameters updated by self-aware attentional backpropagation. We apply PUB-SalNet for analysis of 2D simulated and real images and achieve state-of-the-art performance on simulated biomedical data sets. Furthermore, our proposed PUB-SalNet can be easily extended to 3D images. The experimental results on the 2d and 3d data sets also demonstrate the generalization ability and robustness of our method.

Cellular cryo-Electron Tomography (cryo-ET) provides three-dimensional views of structural and spatial information of various macromolecules in cells in a near-native state. Subtomogram classification is a key step for recognizing and differentiating these macromolecular structures. In recent years, deep learning methods have been developed for high-throughput subtomogram classification tasks; however, conventional supervised deep learning methods cannot recognize macromolecular structural classes that do not exist in the training data. This imposes a major weakness since most native macromolecular structures in cells are unknown and consequently, cannot be included in the training data. Therefore, open set learning which can recognize unknown macromolecular structures is necessary for boosting the power of automatic subtomogram classification. In this paper, we propose a method called Margin-based Loss for Unsupervised Domain Alignment (MLUDA) for open set recognition problems where only a few categories of interest are shared between cross-domain data. Through extensive experiments, we demonstrate that MLUDA performs well at cross-domain open-set classification on both public datasets and medical imaging datasets. So our method is of practical importance.

Cryo-Electron Tomography (cryo-ET) is an emerging 3D imaging technique which shows great potentials in structural biology research. One of the main challenges is to perform classification of macromolecules captured by cryo-ET. Recent efforts exploit deep learning to address this challenge. However, training reliable deep models usually requires a huge amount of labeled data in supervised fashion. Annotating cryo-ET data is arguably very expensive. Deep Active Learning (DAL) can be used to reduce labeling cost while not sacrificing the task performance too much. Nevertheless, most existing methods resort to auxiliary models or complex fashions (e.g. adversarial learning) for uncertainty estimation, the core of DAL. These models need to be highly customized for cryo-ET tasks which require 3D networks, and extra efforts are also indispensable for tuning these models, rendering a difficulty of deployment on cryo-ET tasks. To address these challenges, we propose a novel metric for data selection in DAL, which can also be leveraged as a regularizer of the empirical loss, further boosting the task model. We demonstrate the superiority of our method via extensive experiments on both simulated and real cryo-ET datasets. Our source Code and Appendix can be found at this URL.

The revolutionary technique cryoelectron tomography (cryo-ET) enables imaging of cellular structure and organization in a near-native environment at submolecular resolution, which is vital to subsequent data analysis and modeling. The conventional structure detection process first reconstructs the three-dimensional (3D) tomogram from a series of two-dimensional (2D) projections and then directly detects subcellular components found within the tomogram. However, this process is challenging due to potential structural information loss during the tomographic reconstruction and the limited scope of existing methods since most major state-of-the-art object detection methods are designed for 2D rather than 3D images. Therefore, in this article, as an alternative approach to complement the conventional process, we propose a novel 2D-to-3D framework that detects structures within 2D projection images before reconstructing the results back to 3D. We implemented the proposed framework as three specific algorithms for three individual tasks: semantic segmentation, edge detection, and object localization. As experimental validation of the 2D-to-3D framework for cryo-ET data, we applied the algorithms to the segmentation of mitochondrial calcium phosphate granules, detection of spherical edges, and localization of mitochondria. Quantitative and qualitative results show better performance for prediction tasks of segmentation on the 2D projections and promising performance on object localization and edge detection, paving the way for future studies in the exploration of cryo-ET for in situ structural biology.

Motivation

Cryo-Electron Tomography (cryo-ET) is a 3D bioimaging tool that visualizes the structural and spatial organization of macromolecules at a near-native state in single cells, which has broad applications in life science. However, the systematic structural recognition and recovery of macromolecules captured by cryo-ET are difficult due to high structural complexity and imaging limits. Deep learning-based subtomogram classification has played critical roles for such tasks. As supervised approaches, however, their performance relies on sufficient and laborious annotation on a large training dataset.

Results

To alleviate this major labeling burden, we proposed a Hybrid Active Learning (HAL) framework for querying subtomograms for labeling from a large unlabeled subtomogram pool. Firstly, HAL adopts uncertainty sampling to select the subtomograms that have the most uncertain predictions. This strategy enforces the model to be aware of the inductive bias during classification and subtomogram selection, which satisfies the discriminativeness principle in AL literature. Moreover, to mitigate the sampling bias caused by such strategy, a discriminator is introduced to judge if a certain subtomogram is labeled or unlabeled and subsequently the model queries the subtomogram that have higher probabilities to be unlabeled. Such query strategy encourages to match the data distribution between the labeled and unlabeled subtomogram samples, which essentially encodes the representativeness criterion into the subtomogram selection process. Additionally, HAL introduces a subset sampling strategy to improve the diversity of the query set, so that the information overlap is decreased between the queried batches and the algorithmic efficiency is improved. Our experiments on subtomogram classification tasks using both simulated and real data demonstrate that we can achieve comparable testing performance (on average only 3% accuracy drop) by using less than 30% of the labeled subtomograms, which shows a very promising result for subtomogram classification task with limited labeling resources.

Availability and implementation

https://github.com/xulabs/aitom.

Supplementary information

Supplementary data are available at Bioinformatics online.

Cryo-electron tomography (cryo-ET) provides 3D visualization of subcellular components in the near-native state and at sub-molecular resolutions in single cells, demonstrating an increasingly important role in structural biology in situ. However, systematic recognition and recovery of macromolecular structures in cryo-ET data remain challenging as a result of low signal-to-noise ratio (SNR), small sizes of macromolecules, and high complexity of the cellular environment. Subtomogram structural classification is an essential step for such task. Although acquisition of large amounts of subtomograms is no longer an obstacle due to advances in automation of data collection, obtaining the same number of structural labels is both computation and labor intensive. On the other hand, existing deep learning based supervised classification approaches are highly demanding on labeled data and have limited ability to learn about new structures rapidly from data containing very few labels of such new structures. In this work, we propose a novel approach for subtomogram classification based on few-shot learning. With our approach, classification of unseen structures in the training data can be conducted given few labeled samples in test data through instance embedding. Experiments were performed on both simulated and real datasets. Our experimental results show that we can make inference on new structures given only five labeled samples for each class with a competitive accuracy (> 0.86 on the simulated dataset with SNR = 0.1), or even one sample with an accuracy of 0.7644. The results on real datasets are also promising with accuracy > 0.9 on both conditions and even up to 1 on one of the real datasets. Our approach achieves significant improvement compared with the baseline method and has strong capabilities of generalizing to other cellular components.

Model organisms are instrumental substitutes for human studies to expedite basic, translational, and clinical research. Despite their indispensable role in mechanistic investigation and drug development, molecular congruence of animal models to humans has long been questioned and debated. Little effort has been made for an objective quantification and mechanistic exploration of a model organisms resemblance to humans in terms of molecular response under disease or drug treatment. We hereby propose a framework, namely Congruence Analysis for Model Organisms (CAMO), for transcriptomic response analysis by developing threshold-free differential expression analysis, quantitative concordance/discordance scores incorporating data variabilities, pathway-centric downstream investigation, knowledge retrieval by text mining, and topological gene module detection for hypothesis generation. Instead of a genome-wide vague and dichotomous answer of poorly or greatly mimicking humans, CAMO assists researchers to numerically quantify congruence, to dissect true cross-species differences from unwanted biological or cohort variabilities, and to visually identify molecular mechanisms and pathway subnetworks that are best or least mimicked by model organisms, which altogether provides foundations for hypothesis generation and subsequent translational decisions.