Supplementary data from: Critical role of LdZIP7 in enhancing cadmium tolerance of Lymantria dispar larvae: Functional identification at both the individual and cellular levels

Wang, Ying 1 ; Wang, Ruiqi1; Jiang, Hong1; Tan, Mingtao1; Zhang, Aoying1; He, Yubin1; Yue, Fusen1; Yan, Shanchun1; Jiang, Dun1

Research facility: Northeast Forestry University

Published Jul 07, 2025 on Dryad. https://doi.org/10.5061/dryad.j0zpc86sq

Data files

Jul 07, 2025 version files 1.48 MB

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Abstract

Heavy metals are prevalent environmental pollutants in habitats of phytophagous insects. This study investigates the tolerance of Lymantria dispar larvae to cadmium (Cd) and the associated mechanisms involving the ZIP family. Cd stress reduced larval body weight and extended the development duration without causing significant mortality. A significantly up-regulated expression of apoptosis genes Caspase-1, Caspase-3, and Caspase-7 was observed in Cd-exposed larvae. In Cd-exposed larvae, expression of the apoptosis-inhibitory factor Bcl-2 in the mitochondrial pathway decreased, while apoptosis-inducing factors Bax and AIFM increased. Cd stress markedly elevated the expression of CHOP and Bip, key genes in the endoplasmic reticulum stress pathway. Among the ZIP family genes, LdZIP7 showed the highest up-regulation in response to Cd treatment. Silencing LdZIP7 intensified the negative impacts of Cd stress on L. dispar larvae and significantly reduced the tolerance of L. dispar larvae to Cd. The main manifestations were a further significant decrease in larval body weight, a further significant extension of developmental duration, and the further activation of the mitochondrial pathway and the endoplasmic reticulum stress pathway-triggered apoptosis in Cd-treated larvae. At the Sf9 cell level, LdZIP7 predominantly localizes in the nuclear membrane and cell membrane. Overexpression of LdZIP7 mitigates Cd-induced cytotoxicity by inhibiting the Ca²⁺-MPTP opening degree-mitochondrial membrane potential-apoptosis pathway. Overall, LdZIP7 plays a pivotal role in alleviating the biotoxic effects of Cd and is a significant regulatory gene for Cd tolerance in L. dispar larvae.

We have submitted our raw data (Data.xls) and a folder of unmodified fluorescence micrograph images (Figure).

Variable Descriptions

Data.xls

C: control group
LCd: Low Cd concentration treatment group
HCd: High Cd concentration treatment group
Cd+LdZIP7: Sf9 cells transfected with the recombinant plasmid pAC5.1b-EGFP-2/LdZIP7 served as the experimental subjects
Cd+EGFP: Untreated Sf9 cells and those transfected with EGFP were utilized as negative controls
Figure 1A: The weight of the larvae at different Cd concentrations (g)
Figure 1B: The developmental duration of the larvae under different Cd concentrations (h)
Figure 1C: The mortality rate of the larvae under different Cd concentrations (%)
Figure 2: The expression level of the ZIP gene family genes in the midgut of larvae
Figure S1: The gene expression level after silencing LdZIP7
Figure 3A: The weight of the 4th instar larvae in different treatment groups after silencing LdZIP7 (g)
Figure 3B: The weight of the 5th instar larvae in different treatment groups after silencing LdZIP7 (g)
Figure 3C: The developmental duration of the larvae in different treatment groups after silencing LdZIP7 (h)
Figure 3D: The mortality rate of the larvae in different treatment groups after silencing LdZIP7 (%)
Figure 4: Subcellular localization of LdZIP7 protein in Sf9 cells
Figure S2: Relative expression levels of growth regulatory genes in Cd-treated larvae after LdZIP7 silencing
Figure S3: Relative expression levels of apoptosis indicator genes in Cd-treated larvae after LdZIP7 silencing
Figure S4: Relative expression levels of mitochondrial apoptosis pathway genes in Cd-treated larvae after LdZIP7 silencing
Figure S5: Relative expression levels of endoplasmic reticulum stress pathway genes in Cd-treated larvae after LdZIP7 silencing
Figure S7: The survival rate of Sf9 cells under Cd stress
Figure 5A: Cell viability of different treatment groups (%)
Figure 5B: ROS level of different treatment groups (%)
Figure 5C: Fluorescence images of ROS in different groups
Figure 6A: Fluorescence images of Ca2+ in different groups
Figure 6B: Diagram of Sf9 cell apoptosis in different treatment groups
Figure 6C: Analysis of Ca2+ fluorescence intensity of different treatment groups (%)
Figure 6D: Apoptosis rate of different treatment groups (%)
Figure 7A: The red/green fluorescence representing the MMP
Figure 7B: The green fluorescence representing the MPTP
Figure 7C: Analysis of MMP of different treatment groups (%)
Figure 7D: Analysis of MPTP of different treatment groups (%)

Code/Software

Throughout the entire project, the following tasks were carried out using BioRad, ImageJ, Prism, and SPSS software, respectively: 1) Data collection, 2) data analysis, 3) graph creation, and 4) providing annotations.