Export stoichiometry and contribution of copepod faecal pellets to vertical flux of particulate organic carbon, nitrogen and phosphorus

Sinking of particles is a key mechanism in the transport of organic matter from the ocean’s productive surface layer to the deep sea and sediments, but also constitutes a loss of carbon and growth-limiting nutrients to the pelagic food web. Knowledge on export of particulate phosphorus is limited, particularly in high-latitude regions, in spite of its role as a co-limiting factor in many marine systems. We therefore investigated suspended concentrations and vertical export (by means of sediment traps) of particulate organic carbon (POC), nitrogen (PON) and total particulate phosphorus (TPP) at 3 contrasting sites in the Fram Strait and Barents Sea opening and quantified the contribution of zooplankton faecal pellets to POC, PON and TPP export. The TPP fluxes are the first to be reported from this region and probably the first from the entire Arctic Ocean. The suspended and exported C:N ratios were close to the Redfield ratio and did not differ significantly from each other (mean atomic ratios of 6.1 and 6.3, respectively). The mean C:P of suspended particles (91) was below Redfield, whereas the exported mean (117) exceeded Redfield, indicating more efficient retention of phosphorus than of nitrogen in the water column. Copepod faecal pellets had low C:P ratios and contributed a higher proportion to phosphorus export (mean of 17%) than to carbon and nitrogen export (10%). Faecal pellets may therefore be an important loss factor for phosphorus from the water column compared to slower-sinking material, which is retained more efficiently

Pol λ appears to preferentially generate G:T over C:T mismatches and its misincorporation ability is suppressed by RP-A

<p><b>Copyright information:</b></p><p>Taken from "Human replication protein A can suppress the intrinsic mutator phenotype of human DNA polymerase λ"</p><p>Nucleic Acids Research 2006;34(5):1405-1415.</p><p>Published online 6 Mar 2006</p><p>PMCID:PMC1390690.</p><p>© The Author 2006. Published by Oxford University Press. All rights reserved</p> () Misincorporation of dGTP (lanes 1–3) or dCTP (lanes 4–6) was monitored on the 19/40merT template in the presence of 0.1 mM Mn and increasing concentrations of pol λ (25, 100 and 250, corresponding to 0.25, 1 and 2.5 pmol, respectively). Lane 7: control reaction in the presence of 25 nM pol λ and all four dNTPs at 5 µM each. The positions of the differently migrating dGMP- and dCMP-terminated +1 products are shown on the left side of the panel. () Misincorporation of increasing concentrations of dGTP (lanes 1–6) or dCTP (lanes 7–12) were monitored on the 19/40merT template in the presence of 0.1 mM Mn and 25 nM (0.25 pmol) pol λ. The positions of the differently migrating dGMP- and dCMP-terminated +1 products are shown on the left side of the panel. () Incorporation of dTTP or dATP (5 µM each) was measured in the presence of 50 nM (0.5 pmol) polλ, 5′-labelled d(T)/d(A) primer/template, 1 mM Mg and in the absence (lanes 1 and 9) or in the presence of different concentrations of either PCNA (lanes 2, 3, 10 and 11) or RP-A (lanes 4–6, 12–14), or an equimolar combination of both proteins (lanes 7, 8, 15 and 16). () Incorporation of 5 µM dCTP (lanes 1–8) or 10 µM dGTP (lanes 9–16) was monitored on the 5′-labelled 20/40merG primer/template, in the presence 150 nM (1.5 pmol) pol λ, 0.1 mM Mn, and in the absence (lanes 1 and 9) or in the presence of 0.5 (lanes 2, 10) and 2 pmol (lanes 3, 11) of PCNA, or 0.2 (lanes 4, 12), 0.5 (lanes 5, 13) and 2 pmol (lanes 6, 14) of RP-A, or a combination of 0.2 (lanes 7, 15) or 2 pmol (lanes 15, 16) of PCNA and RP-A in equimolar ratios. The positions of the differently migrating dCMP- and dGMP-terminated primers are indicated on the left side of the panel with an asterisc

Proteins and substrates used in reconstitution of long patch base excision repair Recombinant proteins were purified as described in ‘Materials and Methods’ and separated on a 8–20% gradient SDS-PAGE gel, and stained with Coomassie Blue

<p><b>Copyright information:</b></p><p>Taken from "The checkpoint clamp, Rad9-Rad1-Hus1 complex, preferentially stimulates the activity of apurinic/apyrimidinic endonuclease 1 and DNA polymerase β in long patch base excision repair"</p><p></p><p>Nucleic Acids Research 2007;35(8):2596-2608.</p><p>Published online 10 Apr 2007</p><p>PMCID:PMC1885638.</p><p>© 2007 The Author(s)</p> Lane 1: molecular weight markers; lane 2: APE 1 (2 μg); lane 3: Pol β (2 μg); lane 4: Fen 1 (2 μg); lane 5: Lig I (2 μg); lane 6: 9-1-1 complex (6 μg). Schematic representation of the P-5′-labeled oligonucleotide substrates used in the study: a 100 bp duplex oligonucleotide containing a THF moiety at the position 43 was used for repair reactions, the ends of the substrate were either free (unblocked substrate) or blocked with a biotin at each end (blocked substrate); a 100 bp duplex oligonucleotide with a 1 nucleotide gap at the same position was used for the Pol β assay; with a nick for the Lig I assay and with a 10 nucleotide flap for the Fen 1 reactio

The 9-1-1 complex specifically stimulates the endonuclease activity of APE 1

<p><b>Copyright information:</b></p><p>Taken from "The checkpoint clamp, Rad9-Rad1-Hus1 complex, preferentially stimulates the activity of apurinic/apyrimidinic endonuclease 1 and DNA polymerase β in long patch base excision repair"</p><p></p><p>Nucleic Acids Research 2007;35(8):2596-2608.</p><p>Published online 10 Apr 2007</p><p>PMCID:PMC1885638.</p><p>© 2007 The Author(s)</p> The APE 1 incision assay was performed as described in ‘Materials and Methods’. Reactions were stopped by adding an equal volume of formamide-dye solution and products were analyzed on a 10% denaturing polyacrylamide gel. An APE 1 reaction mixture (10 μl) contained (besides all components described in ‘Materials and Methods’) P-5′-labeled 100 bp duplex oligonucleotide (50 fmol see B), APE 1 (2 fmol). Reactions were incubated for 20 min at 37°C with the indicated amounts of the 9-1-1 complex. As A but with the blocked substrate (50 fmol). Quantification of the stimulation of APE 1 endonuclease cleavage by the 9-1-1 complex on the substrate with free ends (closed circles) and with the ends blocked with biotin (open circles). The values represent the mean of three independent experiments. The error bars correspond to the standard error of the mean

Fine-tuning of the different enzymes in the long patch base excision repair The LP-BER reaction was performed as described in ‘Materials and Methods’

<p><b>Copyright information:</b></p><p>Taken from "The checkpoint clamp, Rad9-Rad1-Hus1 complex, preferentially stimulates the activity of apurinic/apyrimidinic endonuclease 1 and DNA polymerase β in long patch base excision repair"</p><p></p><p>Nucleic Acids Research 2007;35(8):2596-2608.</p><p>Published online 10 Apr 2007</p><p>PMCID:PMC1885638.</p><p>© 2007 The Author(s)</p> All reactions were stopped by adding an equal volume of formamide-dye solution and products were analyzed on a 10% denaturing polyacrylamide gel. The reaction mixtures (10 μl) contained (besides all components described in ‘Materials and Methods’) unlabeled 100 bp duplex oligonuleotide (50 fmol), [α-P] dTTP (2.5 μCi), Pol β (64 fmol), Fen 1 (93 fmol), Lig I (245 fmol) and the indicated amounts of APE 1. Reactions were incubated for 20 min at 37°C. The reaction mixtures (10 μl) contained (besides all components described in ‘Materials and Methods’) P-5′-labeled 100 bp duplex oligonucleotide (50 fmol), APE 1 (55 fmol), Fen 1 (93 fmol) and Lig I (245 fmol). Reactions were incubated for 20 min at 37°C with the indicated amounts of Pol β. The reaction mixtures (10 μl) contained (besides all components described in ‘Materials and Methods’) P-5′-labeled 100 bp duplex oligonucleotide (50 fmol), APE 1 (55 fmol), Pol β (64 fmol), Lig I (245 fmol) and increasing amounts of Fen 1. Reactions were incubated for 20 min at 37°C. The reaction mixtures (1 μl) contained (besides all components described in ‘Materials and Methods’) P-5′-labeled 100 bp duplex oligonucleotide (50 fmol), of APE 1 (55 fmol) of Pol β (64 fmol), Fen 1 (93 fmol) and indicated amounts of Lig I. Reactions were incubated for 20 min at 37°C

Enriched association of WRN–FEN-1 in the chromatin fraction after DNA damage

<p><b>Copyright information:</b></p><p>Taken from "The interaction site of Flap Endonuclease-1 with WRN helicase suggests a coordination of WRN and PCNA"</p><p>Nucleic Acids Research 2005;33(21):6769-6781.</p><p>Published online 2 Dec 2005</p><p>PMCID:PMC1301591.</p><p>© The Author 2005. Published by Oxford University Press. All rights reserved</p> () Greater fraction of WRN and FEN-1 is found in the chromatin fraction after exposure of cells to MMC. Subnuclear fractionation was performed as described in Materials and Methods. Western blot detection of WRN and FEN-1 in different subnuclear fractions of HeLa cells treated with or without MMC (0.5 µg/ml). () Enrichment of WRN and FEN-1 in the chromatin fraction is dependent on MMC dose. Western blot detection of WRN and FEN-1 in the chromatin fractions prepared from HeLa cells treated with or without 0.25, 0.5 or 1 µg/ml MMC. Bottom panel shows Histone H4 detected by western blot

The 9-1-1 complex specifically stimulates the endonuclease activity of APE 1 in LP-BER Schematic representation of the substrate used in the LP-BER reaction

<p><b>Copyright information:</b></p><p>Taken from "The checkpoint clamp, Rad9-Rad1-Hus1 complex, preferentially stimulates the activity of apurinic/apyrimidinic endonuclease 1 and DNA polymerase β in long patch base excision repair"</p><p></p><p>Nucleic Acids Research 2007;35(8):2596-2608.</p><p>Published online 10 Apr 2007</p><p>PMCID:PMC1885638.</p><p>© 2007 The Author(s)</p> LP-BER assay was performed as described in ‘Materials and Methods’ The reaction mixtures (10 μl) contained (besides all components described in ‘Materials and Methods’) unlabeled 100 bp duplex oligonucleotide (50 fmol), [α-P] dTTP (2.5 μCi), APE 1 (4 fmol), Pol β (64 fmol), Fen 1 (93 fmol), and Lig I (245 fmol). Reactions were incubated for 20 min at 37°C with the indicated amounts of the 9-1-1 complex, as B but with the indicated amounts of PCNA as B but with the indicated amounts of BSA. Quantification of the stimulation of APE 1 activity in LP-BER by the 9-1-1 complex (circles; PCNA (rectangles) and BSA (triangles). The values represent the mean of three independent experiments. The error bars correspond to the standard error of the mean. Quantification of the stimulation of APE 1 activity in LP-BER by the 9-1-1 complex, on the substrate with free ends (closed circles and with the ends blocked with biotin (open circles). The values represent the mean of three independent experiments. The error bars correspond to the standard error of the mean

The 9-1-1 complex has no effect on the activities of Fen 1 and Lig I in long patch base excision repair

<p><b>Copyright information:</b></p><p>Taken from "The checkpoint clamp, Rad9-Rad1-Hus1 complex, preferentially stimulates the activity of apurinic/apyrimidinic endonuclease 1 and DNA polymerase β in long patch base excision repair"</p><p></p><p>Nucleic Acids Research 2007;35(8):2596-2608.</p><p>Published online 10 Apr 2007</p><p>PMCID:PMC1885638.</p><p>© 2007 The Author(s)</p> The LP-BER reaction was performed as described in ‘Materials and Methods’. Reactions were stopped by adding an equal volume of formamide-dye solution and products were analyzed on a 10% denaturing polyacrylamide gel. The reaction mixtures (10 μl) contained (besides all components described in Materials and Methods) P-5′-labeled 100 bp duplex oligonucleotide (50 fmol), APE 1 (55 fmol), Pol β (64 fmol), Fen 1 (9 fmol), and of Lig I (245 fmol). Reactions were incubated for 20 min at 37°C with the indicated amounts of the 9-1-1 complex. The reaction mixtures (10 μl) contained (besides all components described in Materials and Methods) P-5′-labeled 100 bp duplex oligonucleotide (50 fmol), APE 1 (55 fmol), Pol β (64 fmol), Fen 1 (93 fmol), and Lig I (25 fmol). Reactions were incubated for 20 min at 37°C with the indicated amounts of the 9-1-1 complex. A Fen 1 reaction mixture (10 μl) contained (besides all components described in Materials and Methods) a 10 nucleotide flap substrate (50 fmol, see B) and Fen 1 (25 fmol). Reactions were incubated for 20 min at 37°C with the indicated amounts of the 9-1-1 complex. A Lig I reaction mixture (10 μl) contained (besides all components described in Materials and Methods) a nicked oligonucleotide (50 fmol, see B) and Lig I (0.5 fmol). Reactions were incubated for 20 min at 37°C with the indicated amounts of the 9-1-1 complex

BLM-interacting region of FEN-1 is required for BLM stimulation of FEN-1 endonucleolytic cleavage of 5′ flap substrate

<p><b>Copyright information:</b></p><p>Taken from "The interaction site of Flap Endonuclease-1 with WRN helicase suggests a coordination of WRN and PCNA"</p><p>Nucleic Acids Research 2005;33(21):6769-6781.</p><p>Published online 2 Dec 2005</p><p>PMCID:PMC1301591.</p><p>© The Author 2005. Published by Oxford University Press. All rights reserved</p> Reaction mixtures (20 µl) containing 10 fmol of the 26 nt 5′ flap substrate, the specified concentrations of wild-type FEN-1, FEN-1ΔC or FEN-1ΔP, and BLM were incubated at 37°C for 15 min under standard conditions as described in Materials and Methods. Products were resolved on 20% polyacrylamide denaturing gels. () Phosphorimage of a typical gel is shown. Lane 1, no enzyme; lanes 2, 4 and 6 are wild-type FEN-1, FEN-1ΔP and FEN-1ΔC, respectively; lanes 3, 5 and 7 are wild-type FEN-1, FEN-1ΔP and FEN-1ΔC, respectively, in the presence of BLM; lane 8, BLM alone. () Per cent incision (mean value of at least three independent experiments with SD indicated by error bars). Quantitative data are shown for incision reactions with FEN-1 or FEN-1ΔC alone (open bars) or in the presence of BLM (light gray bars)

WRN-interacting region of FEN-1 is required for WRN stimulation of FEN-1 endonucleolytic cleavage of 5′ flap substrate

<p><b>Copyright information:</b></p><p>Taken from "The interaction site of Flap Endonuclease-1 with WRN helicase suggests a coordination of WRN and PCNA"</p><p>Nucleic Acids Research 2005;33(21):6769-6781.</p><p>Published online 2 Dec 2005</p><p>PMCID:PMC1301591.</p><p>© The Author 2005. Published by Oxford University Press. All rights reserved</p> Reaction mixtures (20 µl) containing 10 fmol of the 26 nt 5′ flap substrate, the specified concentrations of wild-type FEN-1, FEN-1ΔC or FEN-1ΔP, and either WRN () or WRN () as indicated were incubated at 37°C for 15 min under standard conditions as described in Materials and Methods. Products were resolved on 20% polyacrylamide denaturing gels. Phosphorimages of typical gels are shown. For each gel: lane 1, no enzyme; lanes 2, 4 and 6 are wild-type FEN-1, FEN-1ΔP and FEN-1ΔC, respectively; lanes 3, 5 and 7 are wild-type FEN-1, FEN-1ΔP and FEN-1ΔC, respectively, in the presence of WRN (A) or WRN (B); lane 8, WRN (A) or WRN (B) alone. () Per cent incision (mean value of at least three independent experiments with SDs indicated by error bars). Quantitative data are shown for incision reactions with FEN-1, FEN-1ΔP or FEN-1ΔC alone (open bars), in the presence of WRN (light gray bars) or in the presence of WRN (gray bars)