Lamin A upregulation reorganizes the genome during rod photoreceptor degeneration
Neurodegenerative diseases are accompanied by dynamic changes in gene expression, including the upregulation of hallmark stress-responsive genes. While the transcriptional pathways that impart adaptive and maladaptive gene expression signatures have been the focus of intense study, the role of higher order nuclear organization in this process is less clear. Here, we examine the role of the nuclear lamina in genome organization during the degeneration of rod photoreceptors. Two proteins had previously been shown to be necessary and sufficient to tether heterochromatin at the nuclear envelope. The lamin B receptor (Lbr) is expressed during development, but downregulates upon rod differentiation. A second tether is the intermediate filament lamin A (LA), which is not normally expressed in murine rods. Here, we show that in the rd1 model of retinitis pigmentosa, LA ectopically upregulates in rod photoreceptors at the onset of degeneration. LA upregulation correlated with increased heterochromatin tethering at the nuclear periphery in rd1 rods, suggesting that LA reorganizes the nucleus. To determine how heterochromatin tethering affects the genome, we used in vivo electroporation to misexpress LA or Lbr in mature rods in the absence of degeneration, resulting in the restoration of conventional nuclear architecture. Using scRNA-seq, we show that reorganizing the nucleus via LA/Lbr misexpression has relatively minor effects on rod gene expression. Next, using ATAC-seq, we show that LA and Lbr both lead to marked increases in genome accessibility. Novel ATAC-seq peaks tended to be associated with stress-responsive genes. Together, our data reveal that heterochromatin tethers have a global effect on genome accessibility, and suggest that heterochromatin tethering primes the photoreceptor genome to respond to stress. Overall design: After flow cytometric sorting, cells were barcoded with 'anchor' and 'co-anchor' lipid-modified oligonucleotides generously provided by the Zev Gartner lab. Barcode oligonucleotides were purchased from Integrated DNA Technologies as follows. Barcode 1: F: 5'- CCTTGGCACCCGAGAATTCCAGGAGAAGAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA-3'; Barcode 2: F: 5'- CCTTGGCACCCGAGAATTCCACCACAATGAGAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA-3'; Barcode 3: F: 5'- CCTTGGCACCCGAGAATTCCATGAGACCTAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA-3';  Each replicate was incubated with barcode oligonucleotides for 10 minutes. Cells were pelleted and washed 3 times with PBS. Replicates were pooled and processed in a single 10X Genomics Chromium run.