biostudies-literatureOgawa YNational Institute of Neurological Disorders and StrokeNINDS NIH HHSDr Miriam and Sheldon G. Adelson Medical Research Foundationjcs256180https://www.ebi.ac.uk/biostudies/studies/S-EPMC7972316biostudies-literatureUnknown134(6)Ranbp2 (also known as Nup358) is a member of the nucleoporin family, which constitutes the nuclear pore complex. Ranbp2 localizes at the nuclear membrane and was recently reported at the axon initial segment (AIS). However, we show that the anti-Ranbp2 antibody used in previous studies is not specific for Ranbp2. We mapped the antibody binding site to the amino acid sequence KPLQG, which is present in both Ranbp2 and neurofascin (Nfasc), a well-known AIS protein. After silencing neurofascin expression in neurons, the AIS was not stained by the antibody. Surprisingly, an exogenously expressed N-terminal fragment of Ranbp2 localizes at the AIS. We show that this fragment interacts with stable microtubules. Finally, using CRISPR/Cas9 in primary cultured neurons, we inserted an HA-epitope tag Journal of cell scienceEndogenously expressed Ranbp2 is not at the axon initial segment.PMC7972316R01 NS044916R37 NS044916NIH NS044916Ogawa YRasband MNfalseEndogenously expressed Ranbp2 is not at the axon initial segment.Ranbp2 (also known as Nup358) is a member of the nucleoporin family, which constitutes the nuclear pore complex. Ranbp2 localizes at the nuclear membrane and was recently reported at the axon initial segment (AIS). However, we show that the anti-Ranbp2 antibody used in previous studies is not specific for Ranbp2. We mapped the antibody binding site to the amino acid sequence KPLQG, which is present in both Ranbp2 and neurofascin (Nfasc), a well-known AIS protein. After silencing neurofascin expression in neurons, the AIS was not stained by the antibody. Surprisingly, an exogenously expressed N-terminal fragment of Ranbp2 localizes at the AIS. We show that this fragment interacts with stable microtubules. Finally, using CRISPR/Cas9 in primary cultured neurons, we inserted an HA-epitope tag 2021-01-01T00:00:00Z2021 Mar2025-04-04T10:17:25.424Z2025-04-04T10:17:25.424ZS-EPMC79723163353624910.1242/jcs.256180