Mitochondrial protease Lonp1 localizes to the nucleus in response to heat shock

doi:10.21203/rs.3.rs-227270/v1

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Mitochondrial protease Lonp1 localizes to the nucleus in response to heat shock

https://doi.org/10.21203/rs.3.rs-227270/v1

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The coordinated communication between the mitochondria and nucleus is essential for cellular activities. Nonetheless, the pathways involved in this crosstalk are scarcely understood. The protease Lonp1 was previously believed to be exclusively located in the mitochondria, with an important role in mitochondrial morphology, mtDNA maintenance, and cellular metabolism, in both normal and neoplastic cells. However, we recently detected Lonp1 in the nuclear, where as much as 22% of all cellular Lonp1 can be found. Nuclear localization is detectable under all conditions, but the amount is dependent on a response to heat shock (HS). Lonp1 in the nucleus interacts with heat shock factor 1 (HSF1) and modulates the HS response. Furthermore, we found three splicing variants of Lonp1 mRNA that coexist in cells and generate three Lonp1 isoforms (ISO1, ISO2 and ISO3) with differential subcellular distributions. All three isoforms move into the nucleus under HS, and ISO2 is most frequently upregulated in neoplastic cells compared to ISO1 and ISO3. These findings reveal a novel extramitochondrial function for Lonp1 in response to stress.

Molecular Biology

Cell Cycle & Proliferation

Cellular Metabolism

Mitochondria

Lonp1

HSF1

heat shock

SUPPLEMENTARYFIGURESCMLS.pdf
Supplementary Figure 1 Representative confocal microscopy images of SW48, SW480 and HeLa cells after immunostaining with anti-Lonp1 and anti-hMit antibodies. Nuclei were counterstained with DAPI. Supplementary Figure 2 Principal component analysis and unsupervised cluster analysis and of RNAseq data from SW620 cells undergoing heat shock for 1 hour, heat shock for 1h followed by 1h hour of recovery, and cells kept at 37°C, when Lonp1 was silenced or not. Ctrl siScramble: cells transfected with a scrambled siRNA, kept at 37°C. Ctrl siLonp1: cells transfected with siRNA for LONP1, kept at 37°C. HS siScramble: cells transfected with scrambled siRNA, kept at 42°C for 1 hour. HS_siLONP1: cells transfected with LONP1 siRNA, kept at 42°C for 1 hour. Hs +REC siScramble: cells transfected with scrambled siRNA, kept at 42°C for 1 hour and left at 37°C for 1 hour to recover. HS + Rec siLonp1: cells transfected with LONP1 siRNA, kept at 42°C for 1 hour and left at 37°C for 1 hour to recover. Supplementary Figure 3 Representative confocal microscopy images of HeLa co-transfected with constructs bearing Lonp1 isoforms tagged with enhanced green fluorescent protein (eGFP, in green) or mCherry (in red). Nuclei were counterstained with DAPI. Supplementary Figure 4 Histogram showing the levels of Lonp1 isoform-3 (ISO3) in normal tissues (N) or primary solid tumors (T) obtained from TSVdb. Data represent the mean ± SD. Rectum: rectum adenocarcinoma; Colon: colon adenocarcinoma; Lung: lung adenocarcinoma; Cervical: cervical squamous cell carcinoma and endocervical carcinoma; Bladder: bladder urothelial carcinoma; Prostate: prostate adenocarcinoma; Breast: breast invasive carcinoma; Head and neck: head and neck squamous cell carcinoma; Renal: renal clear cell carcinoma. * P<0.05.
Supplementarytable1.docx
Supplementary table 1. Gene Ontology (GO) terms as determined by DAVID in cells under HS in comparison to cells kept at 37°C, either with silenced Lonp1 (SiLonp1) or in cells treated with a scramble siRNA (Scramble). Significantly enriched terms (Benjamini- Hochberg adjusted p value £ 0.05) are in bold.
Supplementarytable2.docx
Supplementary table 2. Reactome pathways found to be enriched in cells under HS in comparison to cells kept at 37°C, either with silenced Lonp1 (SiLonp1) or in cells treated with a scramble siRNA (siScramble). Significantly enriched pathways (adjusted p value £ 0.05) are in bold.
Supplementarytable3.docx
Supplementary table 3. List of the first 25 genes target of HSF1, upregulated in SW620 cells under HS. The FC with respect to control cells, kept at 37°C, is shown for cells treated with siLonp1 or with a scrambled siRNA. The FC ratio between is also shown.
Supplementarytable4.docx
Supplementary table 4. Enriched gene sets from the Reactome collection in SW620 cells when Lonp1 is silenced, in comparison to scramble siRNA, either at 37°C (Ctrl) and at 42°C for 1 hour (HS), as detected with GSEA. Data are ranked based on Normalized enrichment score (NES).

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Mitochondrial protease Lonp1 localizes to the nucleus in response to heat shock

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