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Fig. 7 LEN is essential for injury-dependent lepb induction during heart regeneration. (A) Guide RNA sequences and genomic coordinates of LEN deletion (LENΔ) generated by CRISPR/Cas9. (B) Representative images of PCNA/Mef2 staining quantified in (C). (C) Quantification of adult ventricular CM proliferation indices for 7 dpa wild-type sibling (control) and LENΔ/Δ hearts. n = 10 and 5 for control and LENΔ/Δ hearts. Three sections per heart were used. (D) Representative images of AFOG staining for 30 dpa control and LENΔ/Δ hearts. n = 6 for control and LENΔ/Δ hearts. Three sections per heart were used. (E) Genomic region surrounding LEN. snd1, lepb, si:dkey-31f5.8 and rap1b are induced during heart regeneration. (F) Transcripts Per Kilobase Million (TPM) for lepb and its surrounding genes. (G) RNA-seq of uninjured and 3 dpa wild-type (WT) and LENΔ/Δ hearts. Tracks indicate an absence of lepb expression at 3 dpa in LENΔ/Δ fish, whereas snd1 and si:dkey-31f5.8 transcript levels are increased similarly in both wild type and LENΔ/Δ upon heart injury. (H) RT-qPCR analysis of snd1, lepb, and si:dkey-31f5.8 transcript levels in WT and LENΔ/Δ hearts. lepb is undetectable in 3 dpa injured LENΔ/Δ hearts. unpaired two-tailed t-tests were performed to indicate significance. (n = 8, 5, 5, and 4 for WT uninjured, LENΔ/Δ uninjured, WT 3dpa, and LENΔ/Δ 3dpa, respectively). Data are mean ± s.d. (I, J) lif/m17 is upregulated upon injury in wild-type and LENΔ/Δ hearts. (I) RNA-seq of uninjured and 3 dpa wild-type (WT) and LENΔ/Δ hearts. Tracks indicate lif/m17 transcript level is increased similarly in both wild type and LENΔ/Δ upon heart injury. (J) RT-qPCR analysis of lif/m17transcript level in WT and LENΔ/Δ hearts. Data are mean ± s.d. (K) RNA-seq of uninjured and 3 dpa wild-type (WT) and LENΔ/Δ hearts. Tracks indicate lepa transcript level is increased similarly in both wild type and LENΔ/Δ upon heart injury