Mol. Cells 2020; 43(3): 228-235
Published online February 6, 2020
https://doi.org/10.14348/molcells.2019.0294
© The Korean Society for Molecular and Cellular Biology
Correspondence to : sjeong4@jbnu.ac.kr
The Drosophila transmembrane semaphorin Sema-1a mediates forward and reverse signaling that plays an essential role in motor and central nervous system (CNS) axon pathfinding during embryonic neural development. Previous immunohistochemical analysis revealed that Sema-1a is expressed on most commissural and longitudinal axons in the CNS and five motor nerve branches in the peripheral nervous system (PNS). However, Sema-1a-mediated axon guidance function contributes significantly to both intersegmental nerve b (ISNb) and segmental nerve a (SNa), and slightly to ISNd and SNc, but not to ISN motor axon pathfinding. Here, we uncover three cis-regulatory elements (CREs), R34A03, R32H10, and R33F06, that robustly drove reporter expression in a large subset of neurons in the CNS. In the transgenic lines R34A03 and R32H10 reporter expression was consistently observed on both ISNb and SNa nerve branches, whereas in the line R33F06 reporter expression was irregularly detected on ISNb or SNa nerve branches in small subsets of abdominal hemisegments. Through complementation test with a Sema1a loss-of-function allele, we found that neuronal expression of Sema-1a driven by each of R34A03 and R32H10 restores robustly the CNS and PNS motor axon guidance defects observed in Sema-1a homozygous mutants. However, when wild-type Sema-1a is expressed by R33F06 in Sema-1a mutants, the Sema-1a PNS axon guidance phenotypes are partially rescued while the Sema-1a CNS axon guidance defects are completely rescued. These results suggest that in a redundant manner, the CREs, R34A03, R32H10, and R33F06 govern the Sema-1a expression required for the axon guidance function of Sema-1a during embryonic neural development.
Keywords axon guidance, cis-regulatory element, Drosophila, motor neurons, semaphorin-1a
The
Here, we identified three GAL4 lines,
The
To precisely examine the expression pattern of each GAL4 line in the PNS and CNS, each GAL4 strain was crossed to
The filleted preparation method was used to better visualize and characterize FasII-positive axon guidance phenotypes observed in the CNS and PNS (Jeong, 2017). The detailed assessment of CNS and motor axon guidance defects were performed as described previously (Van Vactor et al., 1993; Yu et al., 1998). The chi-square (χ2) statistic with a 2 × 2 contingency table was used to assess independence between two categorical variables.
To identify CREs that are required to direct Sema-1a expression during embryonic neural development, we took advantage of a collection of GAL4 lines driven by flanking and intronic DNA fragments of
To find out whether
Since
Here, we provide evidence that endogenous Sema-1a expression is controlled by multiple CREs R34A03, R32H10, and R33F06 in a redundant manner during embryonic neural development. First, both ISNb and SNa motor axon guidance defects seen in
It has been reported that there are usually four developmental enhancer elements per protein-coding gene in the
Since transmembrane Sema-1a functions as a ligand (forward signaling) and/or as a receptor (reverse signaling) to mediate axon-axon repulsion at specific choice points in the PNS (Jeong et al., 2012; 2017; Yu et al., 1998), one important question related to Sema-1a expression is which motoneurons express native Sema-1a proteins during embryonic development. Interestingly, we found that the reporter expression patterns of
The authors have no potential conflicts of interest to disclose.
This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korean government (MIST) (NRF-2018R1A2B6008037 and NRF-2017M3C7A1044816). We thank Alex Kolodkin and the Bloomington Drosophila Stock Center for the fly stocks.
Mol. Cells 2020; 43(3): 228-235
Published online March 31, 2020 https://doi.org/10.14348/molcells.2019.0294
Copyright © The Korean Society for Molecular and Cellular Biology.
Young Gi Hong1 , Bongsu Kang1,2
, Seongsoo Lee3
, Youngseok Lee4
, Bong-Gun Ju5
, and Sangyun Jeong1,2,*
1Division of Life Sciences (Molecular Biology Major), Jeonbuk National University, Jeonju 54896, Korea, 2Department of Bioactive Material Sciences and Research Center of Bioactive Materials, Jeonbuk National University, Jeonju 54896, Korea, 3Gwangju Center, Korea Basic Science Institute, Gwangju 61186, Korea, 4Department of Bio and Fermentation Convergence Technology, BK21 PLUS Project, Kookmin University, Seoul 02707, Korea, 5Department of Life Science, Sogang University, Seoul 04107, Korea
Correspondence to:sjeong4@jbnu.ac.kr
The Drosophila transmembrane semaphorin Sema-1a mediates forward and reverse signaling that plays an essential role in motor and central nervous system (CNS) axon pathfinding during embryonic neural development. Previous immunohistochemical analysis revealed that Sema-1a is expressed on most commissural and longitudinal axons in the CNS and five motor nerve branches in the peripheral nervous system (PNS). However, Sema-1a-mediated axon guidance function contributes significantly to both intersegmental nerve b (ISNb) and segmental nerve a (SNa), and slightly to ISNd and SNc, but not to ISN motor axon pathfinding. Here, we uncover three cis-regulatory elements (CREs), R34A03, R32H10, and R33F06, that robustly drove reporter expression in a large subset of neurons in the CNS. In the transgenic lines R34A03 and R32H10 reporter expression was consistently observed on both ISNb and SNa nerve branches, whereas in the line R33F06 reporter expression was irregularly detected on ISNb or SNa nerve branches in small subsets of abdominal hemisegments. Through complementation test with a Sema1a loss-of-function allele, we found that neuronal expression of Sema-1a driven by each of R34A03 and R32H10 restores robustly the CNS and PNS motor axon guidance defects observed in Sema-1a homozygous mutants. However, when wild-type Sema-1a is expressed by R33F06 in Sema-1a mutants, the Sema-1a PNS axon guidance phenotypes are partially rescued while the Sema-1a CNS axon guidance defects are completely rescued. These results suggest that in a redundant manner, the CREs, R34A03, R32H10, and R33F06 govern the Sema-1a expression required for the axon guidance function of Sema-1a during embryonic neural development.
Keywords: axon guidance, cis-regulatory element, Drosophila, motor neurons, semaphorin-1a
The
Here, we identified three GAL4 lines,
The
To precisely examine the expression pattern of each GAL4 line in the PNS and CNS, each GAL4 strain was crossed to
The filleted preparation method was used to better visualize and characterize FasII-positive axon guidance phenotypes observed in the CNS and PNS (Jeong, 2017). The detailed assessment of CNS and motor axon guidance defects were performed as described previously (Van Vactor et al., 1993; Yu et al., 1998). The chi-square (χ2) statistic with a 2 × 2 contingency table was used to assess independence between two categorical variables.
To identify CREs that are required to direct Sema-1a expression during embryonic neural development, we took advantage of a collection of GAL4 lines driven by flanking and intronic DNA fragments of
To find out whether
Since
Here, we provide evidence that endogenous Sema-1a expression is controlled by multiple CREs R34A03, R32H10, and R33F06 in a redundant manner during embryonic neural development. First, both ISNb and SNa motor axon guidance defects seen in
It has been reported that there are usually four developmental enhancer elements per protein-coding gene in the
Since transmembrane Sema-1a functions as a ligand (forward signaling) and/or as a receptor (reverse signaling) to mediate axon-axon repulsion at specific choice points in the PNS (Jeong et al., 2012; 2017; Yu et al., 1998), one important question related to Sema-1a expression is which motoneurons express native Sema-1a proteins during embryonic development. Interestingly, we found that the reporter expression patterns of
The authors have no potential conflicts of interest to disclose.
This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korean government (MIST) (NRF-2018R1A2B6008037 and NRF-2017M3C7A1044816). We thank Alex Kolodkin and the Bloomington Drosophila Stock Center for the fly stocks.
Sangyun Jeong
Mol. Cells 2021; 44(8): 549-556 https://doi.org/10.14348/molcells.2021.0129Boyoon Choi, Hyeyoung Kim, Jungim Jang, Sihyeon Park, and Hosung Jung
Mol. Cells 2022; 45(11): 846-854 https://doi.org/10.14348/molcells.2022.0081Nari Hong and Yoonkey Nam
Mol. Cells 2022; 45(2): 76-83 https://doi.org/10.14348/molcells.2022.2023