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. 2009 Feb 15;23(4):433–438. doi: 10.1101/gad.1761509

Figure 2.

Figure 2.

GLD-2 is a 3′ adenylase for cytoplasmic ncRNAs. (A) 3′-End sequence of human 7SL RNA gene (NR_002715) and its transcribed sequences with 3′-terminal variations. Cleavage sites of RNase T1 are shown by arrowheads. The five different 3′-terminal fragments are colored yellow (UCUCU), orange (UCUCUu), red (UCUCUuu), light blue (UCUCUa), and blue (UCUCUua). Lowercase letters (u or a) stand for additional nucleosides (uridine or adenosine) attached at the 3′ ends after transcription. 3′-Terminal fragments bearing 2′,3′-cyclic phosphates are not shown. (B) Composition and ratio of 3′ termini of 7SL RNAs isolated from Huh7 cells when each indicated nucleotidyl-transferase was knocked down. The steady-state level of each mRNA in the shRNA treated cells is shown in parentheses. The relative abundance of 3′-terminal fragments was calculated from the intensity of the mass chromatogram for each fragment. The color code of each fragment is the same as in A. (C) Mass chromatograms of doubly charged ions of five different 3′-terminal fragments of 7SL RNAs from Huh7 cells transfected (right panel) or not transfected (left panel) with an shRNA targeting to hGLD-2: UCUCU (m/z 732.00–732.75, yellow), UCUCUu (m/z 885.00–885.75, orange), UCUCUuu (m/z 1038.00–1038.75, red), UCUCUa (m/z 896.50–897.25, light blue), and UCUCUua (m/z 1049.50–1050.25, blue). (D) Mass chromatograms of doubly charged ions of four different 3′-terminal fragments of 7SL RNAs from livers of mGLD-2+/− mice (left panel) or mGLD-2−/− mice (right panel). The color code and m/z value of each fragment are the same as in A–C. (E) Mass chromatograms of the [M-8H]8−, [M-9H]9−, and [M-10H]10− ions of each miR-122 variant [21-nt (MW 6847.84, green), 22-nt (MW 7192.88, black), 23-nt (3′-U) (MW 7498.91, red), and 23-nt (3′-A) (MW 7521.94, blue)] from Huh7 cells transfected (right panel) or not transfected (left panel) with an shRNA targeting hGLD-2. (F) Mass chromatograms of the [M-8H]8−, [M-9H]9−, and [M-10H]10− ions of each miR-122 variant from livers ofmGLD-2+/− mice (left panel) or mGLD-2−/− mice (right panel). The color code and MW value of each variant are the same as in E.