Hongfei Ji

All Publications

• Functional analysis of conserved C. elegans bHLH family members uncovers lifespan control by a peptidergic hub neuron. PLoS biology Aguilar, G. R., Vidal, B., Ji, H., Evenblij, J., Liao, C. P., Ji, H., Valperga, G., Fang-Yen, C., Hobert, O. 2025; 23 (1): e3002979

  Abstract

  Throughout the animal kingdom, several members of the basic helix-loop-helix (bHLH) family act as proneural genes during early steps of nervous system development. Roles of bHLH genes in specifying terminal differentiation of postmitotic neurons have been less extensively studied. We analyze here the function of 5 Caenorhabditis elegans bHLH genes, falling into 3 phylogenetically conserved subfamilies, which are continuously expressed in a very small number of postmitotic neurons in the central nervous system. We show (a) that 2 orthologs of the vertebrate bHLHe22/e23 genes, called hlh-17 and hlh-32, function redundantly to specify the identity of a single head interneuron class (AUA), as well as an individual motor neuron (VB2); (b) that the PTF1a ortholog hlh-13 acts as a terminal selector to control terminal differentiation and function of the sole octopaminergic neuron class in C. elegans, RIC; and (c) that the NHLH1/2 ortholog hlh-15 controls terminal differentiation and function of the peptidergic AVK head interneuron class, a known neuropeptidergic signaling hub in the animal. Strikingly, through null mutant analysis and cell-specific rescue experiments, we find that loss of hlh-15/NHLH in the peptidergic AVK neurons and the resulting abrogation of neuropeptide secretion from these neurons causes a substantially extended lifespan of the animal, which we propose to be akin to hypothalamic control of lifespan in vertebrates. Our functional analysis reveals themes of bHLH gene function during terminal differentiation that are complementary to the earlier lineage specification roles of other bHLH family members. However, such late functions are much more sparsely employed by members of the bHLH transcription factor family, compared to the function of the much more broadly employed homeodomain transcription factor family.

  View details for DOI 10.1371/journal.pbio.3002979

  View details for PubMedID 39761329

  View details for PubMedCentralID PMC11703107

• Automated multimodal imaging of Caenorhabditis elegans behavior in multi-well plates. Genetics Ji, H., Chen, D., Fang-Yen, C. 2024; 228 (4)

  Abstract

  Assays of behavior in model organisms play an important role in genetic screens, drug testing, and the elucidation of gene-behavior relationships. We have developed an automated, high-throughput imaging and analysis method for assaying behaviors of the nematode C. elegans. We use high-resolution optical imaging to longitudinally record the behaviors of 96 animals at a time in multi-well plates, and computer vision software to quantify the animals' locomotor activity, behavioral states, and egg laying events. To demonstrate the capabilities of our system we used it to examine the role of serotonin in C. elegans behavior. We found that egg-laying events are preceded by a period of reduced locomotion, and that this decline in movement requires serotonin signaling. In addition, we identified novel roles of serotonin receptors SER-1 and SER-7 in regulating the effects of serotonin on egg laying across roaming, dwelling, and quiescent locomotor states. Our system will be useful for performing genetic or chemical screens for modulators of behavior.

  View details for DOI 10.1093/genetics/iyae158

  View details for PubMedID 39358843

  View details for PubMedCentralID PMC11631399

• Segmentation-free measurement of locomotor frequency in Caenorhabditis elegans using image invariants. G3 (Bethesda, Md.) Ji, H., Chen, D., Fang-Yen, C. 2024

  Abstract

  An animal's locomotor rate is an important indicator of its motility. In studies of the nematode C. elegans, assays of the frequency of body bending waves have often been used to discern the effects of mutations, drugs, or aging. Traditional manual methods for measuring locomotor frequency are low in throughput and subject to human error. Most current automated methods depend on image segmentation, which requires high image quality and is prone to errors. Here, we describe an algorithm for automated estimation of C. elegans locomotor frequency using image invariants, i.e., shape-based parameters that are independent of object translation, rotation, and scaling. For each video frame, the method calculates a combination of 8 Hu's moment invariants and a set of Maximally Stable Extremal Regions (MSER) invariants. The algorithm then calculates the locomotor frequency by computing the autocorrelation of the time sequence of the invariant ensemble. Results of our method show excellent agreement with manual or segmentation-based results over a wide range of frequencies. We show that compared to a segmentation-based method that analyzes a worm's shape and a method based on video covariance, our technique is more robust to low image quality and background noise. We demonstrate the system's capabilities by testing the effects of serotonin and serotonin pathway mutations on C. elegans locomotor frequency.

  View details for DOI 10.1093/g3journal/jkae170

  View details for PubMedID 39056257

• Functional analysis of conserved C. elegans bHLH family members uncovers lifespan control by a peptidergic hub neuron. bioRxiv : the preprint server for biology Aguilar, G. R., Vidal, B., Ji, H., Evenblij, J., Ji, H., Valperga, G., Liao, C. P., Fang-Yen, C., Hobert, O. 2024

  Abstract

  Throughout the animal kingdom, several members of the basic helix-loop-helix (bHLH) family act as proneural genes during early steps of nervous system development. Roles of bHLH genes in specifying terminal differentiation of postmitotic neurons have been less extensively studied. We analyze here the function of five C. elegans bHLH genes, falling into three phylogenetically conserved subfamilies, which are continuously expressed in a very small number of postmitotic neurons in the central nervous system. We show (a) that two orthologs of the vertebrate bHLHb4/b5 genes, called hlh-17 and hlh-32, function redundantly to specify the identity of a single head interneuron (AUA), as well as an individual motor neuron (VB2), (b) that the PTF1a ortholog hlh-13 acts as a terminal selector to control terminal differentiation and function of the sole octopaminergic neuron class in C. elegans, RIC, and (c) that the NHLH1/2 ortholog hlh-15 controls terminal differentiation and function of the peptidergic AVK head interneuron class, a known neuropeptidergic signaling hub in the animal. Strikingly, through null mutant analysis and cell-specific rescue experiments, we find that loss of hlh-15/NHLH in the peptidergic AVK neurons and the resulting abrogation of neuropeptide secretion causes a substantially expanded lifespan of the animal, revealing an unanticipated impact of a central, peptidergic hub neuron in regulating lifespan, which we propose to be akin to hypothalamic control of lifespan in vertebrates. Taken together, our functional analysis reveals themes of bHLH gene function during terminal differentiation that are complementary to the earlier lineage specification roles of other bHLH family members. However, such late functions are much more sparsely employed by members of the bHLH transcription factor family, compared to the function of the much more broadly employed homeodomain transcription factor family.

  View details for DOI 10.1101/2024.07.12.603289

  View details for PubMedID 39071424

  View details for PubMedCentralID PMC11275782

• Segmentation-free measurement of locomotor frequency in Caenorhabditis elegans using image invariants. bioRxiv : the preprint server for biology Ji, H., Chen, D., Fang-Yen, C. 2024

  Abstract

  An animal's locomotor rate is an important indicator of its motility. In studies of the nematode C. elegans , assays of the frequency of body bending waves have often been used to discern the effects of mutations, drugs, or aging. Traditional manual methods for measuring locomotor frequency are low in throughput and subject to human error. Most current automated methods depend on image segmentation, which requires high image quality and is prone to errors. Here, we describe an algorithm for automated estimation of C. elegans locomotor frequency using image invariants, i.e., shape-based parameters that are independent of object translation, rotation, and scaling. For each video frame, the method calculates a combination of 8 Hu's moment invariants and a set of Maximally Stable Extremal Regions (MSER) invariants. The algorithm then calculates the locomotor frequency by computing the autocorrelation of the time sequence of the invariant ensemble. Results of our method show excellent agreement with manual or segmentation-based results over a wide range of frequencies. We show that compared to the segmentation method that analyzes a worm's shape, our method is more robust to low image quality. We demonstrate the system's capabilities by testing the effects of serotonin and serotonin pathway mutants on locomotor frequency.

  View details for DOI 10.1101/2024.01.16.575892

  View details for PubMedID 38293059

  View details for PubMedCentralID PMC10827210

• A robotic system for automated genetic manipulation and analysis of Caenorhabditis elegans. PNAS nexus Li, Z., Fouad, A. D., Bowlin, P. D., Fan, Y., He, S., Chang, M. C., Du, A., Teng, C., Kassouni, A., Ji, H., Raizen, D. M., Fang-Yen, C. 2023; 2 (7): pgad197

  Abstract

  The nematode Caenorhabditis elegans is one of the most widely studied organisms in biology due to its small size, rapid life cycle, and manipulable genetics. Research with C. elegans depends on labor-intensive and time-consuming manual procedures, imposing a major bottleneck for many studies, especially for those involving large numbers of animals. Here, we describe a general-purpose tool, WormPicker, a robotic system capable of performing complex genetic manipulations and other tasks by imaging, phenotyping, and transferring C. elegans on standard agar media. Our system uses a motorized stage to move an imaging system and a robotic arm over an array of agar plates. Machine vision tools identify animals and assay developmental stage, morphology, sex, expression of fluorescent reporters, and other phenotypes. Based on the results of these assays, the robotic arm selectively transfers individual animals using an electrically self-sterilized wire loop, with the aid of machine vision and electrical capacitance sensing. Automated C. elegans manipulation shows reliability and throughput comparable with standard manual methods. We developed software to enable the system to autonomously carry out complex protocols. To validate the effectiveness and versatility of our methods, we used the system to perform a collection of common C. elegans procedures, including genetic crossing, genetic mapping, and genomic integration of a transgene. Our robotic system will accelerate C. elegans research and open possibilities for performing genetic and pharmacological screens that would be impractical using manual methods.

  View details for DOI 10.1093/pnasnexus/pgad197

  View details for PubMedID 37416871

  View details for PubMedCentralID PMC10321491

• A proprioceptive feedback circuit drives Caenorhabditis elegans locomotor adaptation through dopamine signaling. Proceedings of the National Academy of Sciences of the United States of America Ji, H., Fouad, A. D., Li, Z., Ruba, A., Fang-Yen, C. 2023; 120 (20): e2219341120

  Abstract

  An animal adapts its motor behavior to navigate the external environment. This adaptation depends on proprioception, which provides feedback on an animal's body postures. How proprioception mechanisms interact with motor circuits and contribute to locomotor adaptation remains unclear. Here, we describe and characterize proprioception-mediated homeostatic control of undulatory movement in the roundworm Caenorhabditis elegans. We found that the worm responds to optogenetically or mechanically induced decreases in midbody bending amplitude by increasing its anterior amplitude. Conversely, it responds to increased midbody amplitude by decreasing the anterior amplitude. Using genetics, microfluidic and optogenetic perturbation response analyses, and optical neurophysiology, we elucidated the neural circuit underlying this compensatory postural response. The dopaminergic PDE neurons proprioceptively sense midbody bending and signal to AVK interneurons via the D2-like dopamine receptor DOP-3. The FMRFamide-like neuropeptide FLP-1, released by AVK, regulates SMB head motor neurons to modulate anterior bending. We propose that this homeostatic behavioral control optimizes locomotor efficiency. Our findings demonstrate a mechanism in which proprioception works with dopamine and neuropeptide signaling to mediate motor control, a motif that may be conserved in other animals.

  View details for DOI 10.1073/pnas.2219341120

  View details for PubMedID 37155851

  View details for PubMedCentralID PMC10193984

• A proprioceptive feedback circuit drives Caenorhabditis elegans locomotor adaptation through dopamine signaling. bioRxiv : the preprint server for biology Ji, H., Fouad, A. D., Li, Z., Ruba, A., Fang-Yen, C. 2022

  View details for DOI 10.1101/2022.10.14.512295

• Phase response analyses support a relaxation oscillator model of locomotor rhythm generation in Caenorhabditis elegans. eLife Ji, H., Fouad, A. D., Teng, S., Liu, A., Alvarez-Illera, P., Yao, B., Li, Z., Fang-Yen, C. 2021; 10

  Abstract

  Neural circuits coordinate with muscles and sensory feedback to generate motor behaviors appropriate to an animal's environment. In C. elegans, the mechanisms by which the motor circuit generates undulations and modulates them based on the environment are largely unclear. We quantitatively analyzed C. elegans locomotion during free movement and during transient optogenetic muscle inhibition. Undulatory movements were highly asymmetrical with respect to the duration of bending and unbending during each cycle. Phase response curves induced by brief optogenetic inhibition of head muscles showed gradual increases and rapid decreases as a function of phase at which the perturbation was applied. A relaxation oscillator model based on proprioceptive thresholds that switch the active muscle moment was developed and is shown to quantitatively agree with data from free movement, phase responses, and previous results for gait adaptation to mechanical loadings. Our results suggest a neuromuscular mechanism underlying C. elegans motor pattern generation within a compact circuit.

  View details for DOI 10.7554/eLife.69905

  View details for PubMedID 34569934

  View details for PubMedCentralID PMC8560089

• Distributed rhythm generators underlie Caenorhabditis elegans forward locomotion. eLife Fouad, A. D., Teng, S., Mark, J. R., Liu, A., Alvarez-Illera, P., Ji, H., Du, A., Bhirgoo, P. D., Cornblath, E., Guan, S. A., Fang-Yen, C. 2018; 7

  Abstract

  Coordinated rhythmic movements are ubiquitous in animal behavior. In many organisms, chains of neural oscillators underlie the generation of these rhythms. In C. elegans, locomotor wave generation has been poorly understood; in particular, it is unclear where in the circuit rhythms are generated, and whether there exists more than one such generator. We used optogenetic and ablation experiments to probe the nature of rhythm generation in the locomotor circuit. We found that multiple sections of forward locomotor circuitry are capable of independently generating rhythms. By perturbing different components of the motor circuit, we localize the source of secondary rhythms to cholinergic motor neurons in the midbody. Using rhythmic optogenetic perturbation, we demonstrate bidirectional entrainment of oscillations between different body regions. These results show that, as in many other vertebrates and invertebrates, the C. elegans motor circuit contains multiple oscillators that coordinate activity to generate behavior.

  View details for DOI 10.7554/eLife.29913

  View details for PubMedID 29360037

  View details for PubMedCentralID PMC5780042

• Pan-neuronal imaging in roaming Caenorhabditis elegans. Proceedings of the National Academy of Sciences of the United States of America Venkatachalam, V., Ji, N., Wang, X., Clark, C., Mitchell, J. K., Klein, M., Tabone, C. J., Florman, J., Ji, H., Greenwood, J., Chisholm, A. D., Srinivasan, J., Alkema, M., Zhen, M., Samuel, A. D. 2016; 113 (8): E1082-8

  Abstract

  We present an imaging system for pan-neuronal recording in crawling Caenorhabditis elegans. A spinning disk confocal microscope, modified for automated tracking of the C. elegans head ganglia, simultaneously records the activity and position of ∼80 neurons that coexpress cytoplasmic calcium indicator GCaMP6s and nuclear localized red fluorescent protein at 10 volumes per second. We developed a behavioral analysis algorithm that maps the movements of the head ganglia to the animal's posture and locomotion. Image registration and analysis software automatically assigns an index to each nucleus and calculates the corresponding calcium signal. Neurons with highly stereotyped positions can be associated with unique indexes and subsequently identified using an atlas of the worm nervous system. To test our system, we analyzed the brainwide activity patterns of moving worms subjected to thermosensory inputs. We demonstrate that our setup is able to uncover representations of sensory input and motor output of individual neurons from brainwide dynamics. Our imaging setup and analysis pipeline should facilitate mapping circuits for sensory to motor transformation in transparent behaving animals such as C. elegans and Drosophila larva.

  View details for DOI 10.1073/pnas.1507109113

  View details for PubMedID 26711989

  View details for PubMedCentralID PMC4776525

• Effects of acoustic vibration on the reorientations of C. elegans Chinese Science Bulletin Zhang, Y., Ji, H., Luo, L., Guo, X., Tu, J., Zhang, D. 2016

  View details for DOI 10.1360/N972015-00274

• Low intensity pulse ultrasound stimulate chondrocytes growth in a 3-D alginate scaffold through improved porosity and permeability. Ultrasonics Guo, G., Lu, L., Ji, H., Ma, Y., Dong, R., Tu, J., Guo, X., Qiu, Y., Wu, J., Zhang, D. 2015; 58: 43-52

  Abstract

  A 3-D scaffold culture system has been used to promote in producing functional chondrocytes for repairing damaged cartilage. In the present study, the low intensity pulse ultrasound (LIPUS) (P(-)=0, 0.055, 0.085 and 0.11 MPa) was applied to improve the porosity and permeability of a 3-D alginate scaffold which was beneficial for the nutrition supply and metabolism during cell growth in 3-D alginate scaffold. The porosity and permeability of the scaffold was quantitatively analyzed based on scanning electron microscopy examination and fluorescence image observation. The results suggest that, for the scaffold exposed to LIPUS, its porosity and permeability could be significantly enhanced by the increasing LIPUS amplitude, which might be induced by the microstreaming shear stress generated by ultrasound-driven microbubble oscillations. Furthermore, the assessments of cell proliferation and collagen II expression confirmed that chondrocytes growth could be effectively promoted in 3-D alginate scaffolds treated by LIPUS, because of the improved scaffold porosity and permeability might benefit cell growth space and nutrition supply. It should also be noticed that appropriate LIPUS driving parameters should be adapted to achieve optimized chondrocytes culture effect in 3-D alginate scaffold.

  View details for DOI 10.1016/j.ultras.2014.12.001

  View details for PubMedID 25543661

• Ultrasonic enhancement of the porosity of alginate scaffold ACTA PHYSICA SINICA Lu Lu, Ji Hong-Fei, Guo Ge-Pu, Guo Xia-Sheng, Tu Juan, Qiu Yuan-Yuan, Zhang Dong 2015; 64 (2)

  View details for DOI 10.7498/aps.64.024301

  View details for Web of Science ID 000349455100034