Choline transporter-like proteins 1 and 2 are newly identified plasma membrane and mitochondrial ethanolamine transporters

Abstract

The membrane phospholipids phosphatidylcholine and phosphatidylethanolamine (PE) are synthesized de novo by the CDP-choline and CDP-ethanolamine (Kennedy) pathway, in which the extracellular substrates choline and ethanolamine are transported into the cell, phosphorylated, and coupled with diacylglycerol to form the final phospholipid product. Although multiple transport systems have been established for choline, ethanolamine transport is poorly characterized and there is no single protein assigned a transport function for ethanolamine. The solute carriers 44A (SLC44A) known as choline transporter-like proteins-1 and -2 (CTL1 and CTL2) are choline transporter at the plasma membrane and mitochondria. We report a novel function of CTL1 and CTL2 in ethanolamine transport. Using the lack or the gain of gene function in combination with specific antibodies and transport inhibitors we established two distinct ethanolamine transport systems of a high affinity, mediated by CTL1, and of a low affinity, mediated by CTL2. Both transporters are Na + -independent ethanolamine/H + antiporters. Primary human fibroblasts with separate frameshift mutations in the CTL1 gene (M1= SLC44A1 ΔAsp517 and M2= SLC44A1 ΔSer126 ) are devoid of CTL1 ethanolamine transport but maintain unaffected CTL2 transport. The lack of CTL1 in M2 cells reduced the ethanolamine transport, the flux through the CDP-ethanolamine Kennedy pathway, and PE synthesis. In contrast, overexpression of CTL1 in M2 cells improved ethanolamine transport and PE synthesis. These data firmly establish that CTL1 and CTL2 are the first identified ethanolamine transporters in whole cells and mitochondria, with intrinsic roles in de novo PE synthesis by the Kennedy pathway and intracellular redistribution of ethanolamine.

Organelle-selective click labeling coupled with flow cytometry allows high-throughput CRISPR screening of genes involved in phosphatidylcholine metabolism

“…To confirm this, we first quantified natural choline extracted from cytoplasm using a choline oxidase-mediated coupled enzyme assay, which revealed that the loss of FLVCR1 dramatically reduced the amount of endogenous choline inside cells (by nearly 80%) (Figure 4D). Note that these phenotypes of FLVCR -KO (decrease in PC synthesis and choline uptake) are the same phenotypes associated with major choline transporter inhibition (Taylor et al, 2021). Moreover, we found that the lower PC labeling intensity in FLVCR1 -KO cells was partially restored by re-expression of FLVCR1 and completely rescued by overexpression of SLC5A7 (a high affinity choline transporter specifically expressed in cholinergic neurons) (Okuda et al, 2000); whereas overexpression of PCYT1A had little effect (Figure 4E).…”

Construction of a novel choline metabolism-related signature to predict prognosis, immune landscape, and chemotherapy response in colon adenocarcinoma

“…CHKB and PEMT are the key genes used to construct the choline metabolism-related signature. CHKB, choline kinase beta, is the first reaction enzyme of the Kennedy pathway, which activates the choline metabolic pathway by catalyzing the biosynthesis of phosphatidylethanolamine and phosphatidylethanol (28,29). Oliveira et al found that a novel splicing mutation of CHKB results in a lack of phosphatidylcholine in muscle fibers by whole-exome sequencing technology, which in turn leads to congenital muscular dystrophies (30).…”

Blood–Brain Barrier Solute Carrier Transporters and Motor Neuron Disease

“…In the mitochondria and plasma membrane, CTL1 is a choline /H + antiporter [ 102 ]. The exact role and function of CTL2/Slc44a2 are not well known; however, it is indirectly involved in phosphatidylcholine synthesis [ 103 ]. Choline scarcity affects various processes, including the expression of genes involved in cell differentiation, apoptosis, and proliferation.…”