Biology subjects

Seong, J.

Publications and source records attributed to Seong, J..

5 recordsLinked to original sources

bioRxiv2025

A non-destructive approach to assess the gut microbiome of honey bee (Apis mellifera) queens using fecal samples

Fecal sampling is a widely used, non-invasive method for assessing gut microbiomes across various organisms. However, its suitability for studying the gut microbiome of honey bee (Apis mellifera) queens has not been tested. In this study, we evaluated whether fecal microbiomes accurately reflect gut microbiomes in honey bee queens, offering a potential non-destructive approach for microbiome research. We successfully obtained fecal and gut samples from 21 out of 26 queens. Bacterial communities were analyzed using 16S rRNA amplicon sequencing and qPCR. Our results indicate that queen fecal microbiomes closely resemble gut microbiomes, with no significant differences in alpha diversity and only minor differences in specific bacterial taxa. Beta diversity analyses revealed that within-pair microbiomes (i.e., gut vs. feces from the same queen) were significantly more similar than between-pair comparisons. Additionally, qPCR analyses revealed a strong positive correlation between bacterial abundances in fecal and gut samples, further supporting the use of feces as a proxy for gut microbiome composition. While promising, fecal collection from queens can sometimes be challenging. In our study, we were unable to collect feces from five queens, and these individuals lacked stored fecal material upon inspection of dissected guts. Nonetheless, our findings suggest that fecal sampling can be a useful, non-invasive method for studying honey bee queen microbiomes, enabling longitudinal assessments without compromising colony stability. ImportanceThe gut microbiome plays important roles in honey bee health, including protection against pathogens and stimulation of host physiological pathways. Because gut microbiome analysis involves destructive dissections, most research has focused on workers, given that colonies can have tens of thousands of individuals. This has left a significant gap in microbiome studies of other colony members, such as queens. Given that a queen is the sole reproductive female, her gut microbiome cannot be sampled without compromising colony integrity. In this study, we demonstrate that fecal sampling from honey bee queens provides a reliable, non-destructive alternative for assessing the composition of their gut microbiomes. We found that queen fecal microbiomes closely resemble their gut microbiomes in both taxonomic diversity and composition. This approach enables longitudinal monitoring of queen microbiomes without disrupting colony integrity, offering a valuable tool for honey bee research.

microbiology↗

bioRxiv2022

Precise temporal control of GEVI conformations enables the visualization of charge migration in a fluorescent protein resulting in an improved optical response

Rapid and reproducible optical transitions of a fluorescent protein (FP) can be achieved with a Genetically Encoded Voltage Indicator (GEVI) via manipulation of the membrane potential. These transitions revealed novel effects of internal mutations near the chromophore that would not be detected under steady state conditions. Mutating an internal threonine (T203) affected the speed of the voltage-dependent fluorescence transition suggesting a conformational change inside the protein. These optical transitions also demonstrated interplay between internal and externally oriented sidechains of the {beta}-can structure. Replacing the steric hindrance of a phenylalanine near the chromophore with threonine (F165T) did not alter the resting fluorescence but resulted in a more complex fluorescent transition providing evidence for a flexible chromophore undergoing conformational changes. F165T orientation was influenced by the flanking external amino acids at positions 164 and 166 with 164F/165T/166T exacerbating the complexity of the voltage-dependent transition while 164T/165T/166F reduced the flexibility of the chromophore resembling the transition pattern of the original F165 version. Alphafold predictions reveal a threonine switch with different orientations of the F165T internal side chain depending on the direction of the offset in polarity at external positions 164 and 166. The crystal structures of the pH-sensitive FP, Super Ecliptic pHluorin and two derivatives solved in varying pH conditions also indicate interactions between the external protein surface and the internal environment providing another example of a threonine switch near the chromophore at T203. This ability to orient internal sidechains has led to the development of a novel GEVI that gets brighter upon depolarization of the plasma membrane, works at low light levels, is less susceptible to physiological pH, and provides in vivo signals. These observations affecting fluorescent transitions should also prove valuable to the development of any FP-based biosensor.

neuroscience↗

bioRxiv2021

Differential receptor crosstalk in DRD1-DRD2 heterodimer upon phasic and tonic dopamine signals

In response to phasic and tonic release, dopamine neurotransmission is regulated by its receptor subtypes, mainly dopamine receptor type 1 and 2 (DRD1 and DRD2). These dopamine receptors are known to form a heterodimer, however the receptor crosstalk between DRD1 and DRD2 was only suspected by measuring their downstream signaling products, due to the lack of methodology for selectively detecting individual activity of different dopamine receptors. Here, we develop red DRD1 sensor (R-DRD1) and green DRD2 sensor (G-DRD2) which can specifically monitor the real-time activity of DRD1 and DRD2, and apply these multicolor sensors to directly measure the receptor crosstalk in the DRD1-DRD2 heterodimer. Surprisingly, we discover that DRD1 activation in the heterodimer is inhibited only at micromolar phasic concentration of dopamine, while DRD2 activation is selectively inhibited at nanomolar tonic dopamine level. Differential receptor crosstalk in the DRD1-DRD2 heterodimer further modulates their downstream cAMP level. These data imply a novel function of the DRD1-DRD2 heterodimer at physiological dopamine levels of phasic and tonic release. Our approach utilizing multicolor receptor sensors will be useful to discover novel function of GPCR heterodimers.

neuroscience↗

bioRxiv2021

Optical regulation of endogenous RhoA reveals switching of cellular responses by signal amplitude

How protein signaling networks respond to different input strengths is an important but poorly understood problem in cell biology. For example, the small GTPase RhoA regulates both focal adhesion (FA) growth or disassembly, but whether RhoA serves as a switch selecting between cellular outcomes, or if outcomes are simply modulated by additional factors in the cell, is not clear. Here, we develop a photoswitchable RhoA guanine exchange factor, psRhoGEF, to precisely control endogenous RhoA activity. We also develop a FRET-based biosensor to allow visualization of RhoA activity together with psRhoGEF control. Using these new optical tools, we discover that low levels of RhoA activation preferentially induce FA disassembly in a Src-dependent manner, while high levels induce both FA growth and disassembly in a ROCK-dependent manner. Thus, rheostatic control of RhoA activation with photoswitchable RhoGEF reveals that cells can use signal amplitude to produce multiple responses to a single biochemical signal.

bioengineering↗

bioRxiv2018

Activation of RIG-I-mediated antiviral signaling triggers autophagy through the MAVS-TRAF6-Beclin-1 signaling axis

Autophagy has been implicated in innate immune responses against various intracellular pathogens. Recent studies have reported that autophagy can be triggered by pathogen recognizing sensors, including Toll-like receptors and cyclic guanosine monophosphate-adenosine monophosphate synthase, to participate in innate immunity. In the present study, we examined whether the RIG-I signaling pathway, which detects viral infections by recognizing viral RNA, triggers the autophagic process. The introduction of polyI:C into the cytoplasm, or Sendai virus infection, significantly induced autophagy in normal cells but not in RIG-I-deficient cells. PolyI:C transfection or Sendai virus infection induced autophagy in the cells lacking type-I interferon signaling. This demonstrated that the effect was not due to interferon signaling. RIG-I-mediated autophagy diminished by the deficiency of mitochondrial antiviral signaling protein (MAVS) or tumor necrosis factor receptor-associated factor (TRAF)6, showing that the RIG-I-MAVS-TRAF6 signaling axis was critical for RIG-I-mediated autophagy. We also found that Beclin-1 was translocated to the mitochondria, and it interacted with TRAF6 upon RIG-I activation. Furthermore, Beclin-1 underwent K63-polyubiquitination upon RIG-I activation, and the ubiquitination decreased in TRAF6-deficient cells. This suggests that the RIG-I-MAVS-TRAF6 axis induced K63-linked polyubiquitination of Beclin-1, which has been implicated in triggering autophagy. Collectively, the results of this study show that the recognition of viral infection by RIG-I is capable of inducing autophagy to control viral replication. As deficient autophagy increases the type-I interferon response, the induction of autophagy by the RIG-I pathway might also contribute to preventing an excessive interferon response as a negative-feedback mechanism.\n\nImportanceMammalian cells utilize various innate immune sensors to detect pathogens. Among those sensors, RIG-I recognizes viral RNA to detect intracellular viral replication. Although cells experience diverse physiological changes upon viral infection, studies to understand the role of RIG-I signaling have focused on the induction of type-I interferon. Autophagy is a process that sequesters cytosolic regions and degrades the contents to maintain cellular homeostasis. Autophagy participates in the immune system, and has been known to be triggered by some innate immune sensors, such as TLR4 and cGAS. We demonstrated that autophagy can be triggered by the activation of RIG-I. In addition, we also proved that MAVS-TRAF6 downstream signaling is crucial for the process. Beclin-1, a key molecule in autophagy, is translocated to mitochondria, where it undergoes K63-ubiquitination in a TRAF6-dependent manner upon RIG-I activation. As autophagy negatively regulates RIG-I-mediated signaling, the RIG-I-mediated activation of autophagy may function as a negative-feedback mechanism.

immunology↗