Heme and sensory neuropathy: insights from novel mutations in the heme exporter feline leukemia virus subgroup C receptor 1

Abstract

Hereditary sensory and autonomic neuropathies (HSANs) are a group of clinically and genetically heterogeneous disorders of the peripheral nervous system mainly characterized by impaired nociception and autonomic dysfunction. We previously identified heme metabolism as a novel pathway contributing to sensory neurons maintenance and nociception. Indeed, we reported mutations in the feline leukemia virus subgroup C receptor 1 (FLVCR1) gene in individuals affected by HSAN. FLVCR1 gene encodes for 2 heme export proteins, FLVCR1a (plasma membrane) and FLVCR1b (mitochondria), crucially involved in the regulation of cellular heme homeostasis. Here, we report on 2 additional patients carrying novel biallelic mutations in FLVCR1 translation initiation codon (c.2T>C; p.(Met1Thr) and c.3G>T; p.(Met1Ile)). We overexpressed the c.2T>C; p.(Met1Thr) mutant in human cell lines and we describe its impact on protein structure and function in comparison with other HSAN-related mutations. We found that the mutation interferes with translation in 2 different ways: by lowering levels of translation of wild-type protein and by inducing translation initiation from a downstream in-frame ATG, leading to the production of an N-terminal truncated protein that is retained in the endoplasmic reticulum. The impact of different kinds of mutations on FLVCR1a localization and structure was also described. The identification of novel FLVCR1 mutations in HSAN reinforces the crucial role of heme in sensory neuron maintenance and pain perception. Moreover, our in vitro findings demonstrate that heme export is not completely lost in HSAN patients, thus suggesting the possibility to improve FLVCR1 expression/activity for therapeutic purposes.

Hereditary Ataxia: A Focus on Heme Metabolism and Fe-S Cluster Biogenesis

“…The observation that loss of Flvcr1 is incompatible with life in animal models [36,92,93] strongly suggests that the identified mutations in FLVCR1 did not completely abrogate the expression of the transporter. This statement is also supported by data obtained in cells expressing the FLVCR1 mutations identified in patients with pain insensitivity [89,90].…”

“…Both homozygous missense mutations and compound heterozygous mutations have been described in several families [79][80][81][82][83]. Interestingly, mutations in the same gene have been identified in patients with non-syndromic retinitis pigmentosa [84][85][86][87][88] or in patients with pain insensitivity [89][90][91]. All the FLVCR1 mutations identified to date in PCARP patients are reported in Table 2.…”

The heme synthesis-export system regulates the tricarboxylic acid cycle flux and oxidative phosphorylation

“…To address that point, we down-modulated FLVCR1a in neuroblastoma SH-SY5Y cells (Figures S7A and S7B) and in tumor endothelial cells (BTECs, breast-cancerderived tumor endothelial cells) (Figures S7C and S7D). We chose such models based on previous observations that FLVCR1 inactivation leads to reduced survival/proliferation in the nervous and endothelial system (Chiabrando et al, 2016;Petrillo et al, 2018;Bertino et al, 2019).…”

“…Gene overexpression was performed as described in Bertino et al (2019). Briefly, the FLVCR1a cDNA present in pCMV-SPORT6 vector (MGC human FLVCR1 sequence, clone 4417876, Horizon Discovery, Cambridge, UK, catalog n MHS6278-202757940) and the Myc-tag sequence present in pcDNA3.1(-)/myc-His B (Thermofisher Scientific, Waltham, MA USA, catalog n V85520) were cloned in frame in the pLVX-puro vector, a lentiviral expression vector with constitutively active human cytomegalovirus immediate early promoter and the puromycin resistance gene (Clontech Laboratories Inc. A Takara Bio Company, Mountain View, CA, USA, catalog number 632164), in order to obtain the pLVX-puro FLVCR1a vector.…”

Distinct Mechanisms Account for In Vitro Activation and Sensitization of TRPV1 by the Porphyrin Hemin

“…Saying this, Chiabrando and co-workers recently pledged for an increased attention to the yet poorly examined role of hemin as a relevant player in nociception [ 29 ]. The authors mainly referred to recent findings showing that loss-of-function variants of the gene encoding for the heme export protein FLVCR1 (feline leukemia virus subgroup receptor 1) can cause hereditary sensory and autonomic neuropathies with a loss of pain perception [ 29 , 30 , 31 ]. A likely mechanism for this disorder is an accumulation of intracellular heme and a consecutive oxidative stress triggering neurotoxicity and thus sensory denervation [ 30 ].…”