Abstract
BackgroundThe chromatin remodeler NAP1L1, which is upregulated in small intestinal neuroendocrine neoplasms (NENs), has been implicated in cell cycle progression. As p57Kip2 (CDKN1C), a negative regulator of proliferation and a tumor suppressor, is controlled by members of the NAP1 family, we tested the hypothesis that NAP1L1 may have a mechanistic role in regulating pancreatic NEN proliferation through regulation of p57Kip2.ResultsNAP1L1 silencing (siRNA and shRNA/lipofectamine approach) decreased proliferation through inhibition of mechanistic (mammalian) target of rapamycin pathway proteins and their phosphorylation (p < 0.05) in the pancreatic neuroendocrine neoplasm cell line BON in vitro (p < 0.0001) and resulted in significantly smaller (p < 0.05) and lighter (p < 0.05) tumors in the orthotopic pancreatic NEN mouse model. Methylation of the p57 Kip2 promoter was decreased by NAP1L1 silencing (p < 0.05), and expression of p57Kip2 (transcript and protein) was upregulated. For methylation of the p57 Kip2 promoter, NAP1L1 bound directly to the promoter (−164 to +21, chromatin immunoprecipitation). In 43 pancreatic NEN samples (38 primaries and 5 metastasis), NAP1L1 was over-expressed in metastasis (p < 0.001), expression which was inversely correlated with p57Kip2 (p < 0.01) on mRNA and protein levels. Menin was not differentially expressed.ConclusionNAP1L1 is over-expressed in pancreatic neuroendocrine neoplasm metastases and epigenetically promotes cell proliferation through regulation of p57 Kip2 promoter methylation.
Highlights
The chromatin remodeler nucleosome assembly protein 1-like 1 (NAP1L1), which is upregulated in small intestinal neuroendocrine neoplasms (NENs), has been implicated in cell cycle progression
Since one of the major inherited genes involved in pancreatic NENs is menin, a known growth inhibitor and chromatin remodeler [30,31], we examined its expression to assess whether it was associated with NAP1L1 and p57Kip2
While other NAP1 family members, e.g., NAP1L2, NAP1L3, and NAP1L5 [11,12,13,14] are well known to be neuron-specific and play a role in proliferation [15] and the NAP1 family is associated with transcriptional regulation [8], our current results define a role for NAP1L1 in the transcriptional regulation of pancreatic neuroendocrine neoplasm cell cycle progression
Summary
The chromatin remodeler NAP1L1, which is upregulated in small intestinal neuroendocrine neoplasms (NENs), has been implicated in cell cycle progression. As p57Kip (CDKN1C), a negative regulator of proliferation and a tumor suppressor, is controlled by members of the NAP1 family, we tested the hypothesis that NAP1L1 may have a mechanistic role in regulating pancreatic NEN proliferation through regulation of p57Kip. The epigenome is a potential target in the etiopathology of pNENs. One gene that was not mutated was nucleosome assembly protein 1-like 1 (NAP1L1). NAP1L1 belongs to a family which is thought to be involved in nucleosome assembly and exchange of histone H2A-H2B dimmers [5,6,7], as well as transcriptional regulation and cell cycle progression [8]. NAP1-like proteins have similar activities regarding nucleosome assembling; NAP1L1, displays the highest disassembly activity [16]. Little is known regarding its expression and potential role in the pathogenesis of pNENs
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