A Complex genomic locus drives mtDNA replicase POLG expression to its Disease-Related Nervous System Regions (2017)
Figure 3

Figure 3. LINC00925 regulates POLG expression

The number of distal DNAse I hypersensitive sites (DHSs) of mtDNA maintenance genes and genes in the genomic POLG locus correlating >0.85 with target promoter DHS across 125 cell lines. Abbreviations: TWNK, Twinkle protein; POLG, DNA Polymerase gamma, catalytic subunit; POLG2, DNA Polymerase gamma, accessory subunit; SSBP1, Single stranded DNA binding protein 1; TFAM, mitochondrial transcription factor A; FANCI, Fanconi anemia group I protein; RHCG, Ammonium transporter Rh type C; TICRR, Treslin.

Distribution histogram of DHSs for mtDNA maintenance genes.

Genomic distribution of POLG DHSs. Black arrow shows a cluster upstream from POLG coding region.

Genes surrounding LINC00925.

Conservation of regulatory elements of POLG genomic locus in species: LINC00925, POLG EEs, MIR9-3. Abbreviations: PanTro, Pan troglodytes; MM, Mus musculus; MonDom, Monodelphis domestica; Xen, Xenopus levis. Adapted from https://rvista.dcode.org.

Expression of LINC00925 and POLG in different human tissues and cell types. Quantitative PCR amplification of cDNA. Abbreviations: iPS, induced pluripotent stem cell; SH5Y, neuroblastoma line; HepG2, liver hepatocellular carcinoma line; U2OS bone osteosarcoma line. Error bars indicate standard deviation in 3 technical replicates. AU; arbitrary units.

Polg and long-non-coding RNA Ai854517 (mouse homologue of human LINC00925) correlate tightly in mouse cerebellar development. Time points: E18, postnatal days 0, 3, 6, 9; three mice per time point. Expression calculated as cap analysis of gene expression (CAGE) hits in the transcription start site (CTSS).

Ai854517 and Polg transcripts colocalize in adult mouse brain; in situ hybridization. HC, hippocampus; cerebellar molecular layer (M), Purkinje cell layer (PC), granular cell layer (GC). Scale bars: hippocampus 760 µm, cerebellum 300 µm.

Expression levels of MIR9-3 and Ai854517 correlate in mouse cerebellar development. Time points: E18, postnatal day 0, 3, 6, 9; three mice per time point.. Expression calculated as cap analysis of gene expression (CAGE) hits in the transcription start site (CTSS).

Predicted targets of MIR9. Six targets were predicted by all prediction programs, TargetScan, PicTar, miRDB and PITA: low density lipoprotein receptor adaptor protein-1, LDLRAP1; methylene-tetrahydrofolate-dehydrogenase-2, MTHFD2; follistatin-like-1, FSTL-1, cytokine; capping-Actin Protein of muscle Z-line alpha-subunit 1, CAPZA1, involved in actin polymerization; PR-domain zinc-finger-domain-1, PRDM1, a repressor of beta-interferon expression; PRRX, a homeobox protein regulator of bone development. One cut homeobox 1 and 2 (ONECUT1 and ONECUT2) and nuclear receptor subfamily 2 group E member 1 (NR2E1) are recently discovered MIR9 targets.

RNA expressions of MIR9, NR2E2, LDLDRAP1 and MTHFD2 in HEK293 untransfected controls and in cells transfected with pre-MIR9 or scrambled RNA. Shown is mean with standard error of the mean. Statistical testing was performed using 1-way ANOVA with Dunnet's correction for multiple comparisons. Abbreviations: AU; arbitrary units.