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. 2019 Dec 20;27(6):1998–2013. doi: 10.1038/s41418-019-0483-6

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© The Author(s), under exclusive licence to ADMC Associazione Differenziamento e Morte Cellulare 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — a Ectopically expressed FLAG-hnRNP F or H1 in HeLa cells were immunoprecipitated by FLAG antibody, and the immunoprecipitates were separated by SDS-PAGE and silver-stained. The protein bands were retrieved and analyzed by mass spectrometry. The numbers of unique peptide and unique spectrum of potential binding proteins implicated in telomerase holoenzyme are listed. HeLa cells transiently co-expressing HA-tagged hnRNP F (b) or H1 (c) with FLAG-tagged DKC1, TCAB1 or hTERT, respectively, were harvested and immunoprecipitated (IP) using anti-FLAG antibodies in the absence or in the presence of RNase A treatment. The immunoprecipitates were subjected to western blot analysis for the indicated proteins. Whole cell lysates served as input. d HeLa cell extracts stably overexpressed FLAG-tagged hnRNP F, H1, H2, or TCAB1 incubated with anti-FLAG antibodies and the immunoprecipitates (IP) were used for the TRAP assay to detect co-immunoprecipitated telomerase activity. The Vector served as a negative control, and FLAG-TCAB1 was used as a positive control. IC: Internal Control (36 bp). e Data from d were quantified and graphed. Error bars represent means ± SD (n = 3). Statistical analysis was performed using Student's t test (** P < 0.01, *** P < 0.001). HeLa cells transiently overexpressing FLAG tagged wild-type hnRNP F or its ΔqRRM trunctions f, H1 or its ΔqRRM trunctions g, were performed IP-TRAP to assess telomerase activity. The expressions of FLAG-hnRNP F/H1 and truncations were analyzed by Western blot. Vector served as a negative control. Error bars represent means ± SD (n = 3). Statistical analysis was performed using Student's t test (*P < 0.05, **P < 0.01, ***P < 0.001). h HeLa cells transiently co-expressing FLAG-tagged hnRNP F or H1 with HA-tagged indicated proteins were subjected to IP-TRAP. Error bars represent means ± SD (n = 3). Statistical analysis was performed using Student's t test (**P < 0.01, ***P < 0.001). WCL, whole cell lysate.