Incubation Setup:
Surface water was collected for shipboard incubations in June 2022 aboard the R/V Sikuliaq using a trace metal clean surface pump "towfish" system (Mellett and Buck, 2020). Filtered (<0.2-micrometer (µm), Acropak) seawater from the towfish was homogenized in three acid-cleaned and seawater rinsed 50-liter (L) carboys that were filled round-robin style (Burns et al., 2023). Each carboy was then bubbled overnight with a custom CO2-air mixture to achieve the target pH levels of pH 8.1, 7.6, and 7.1, which was verified with shipboard spectrophotometric pH analyses using the Byrne MICA system (Adornato et al., 2016). Unfiltered surface seawater was then collected and homogenized in a fourth acid-cleaned and seawater-rinsed carboy using the trace metal clean towfish. Trace metal clean polycarbonate incubation bottles were then filled two-thirds with filtered seawater and one-third unfiltered seawater, amended for the nutrient and/or iron treatment, sealed with the caps/threads wrapped in parafilm and electrical tape, and delivered to deckboard flow-through seawater incubators that were covered in screening to mimic surface light levels. Once all incubation bottles were in the incubators, the time-zero sampling of each incubation began. Six incubations were conducted, two each at a coastal upwelling station (Inc 1, 2; 40.112 ºN, 125.56 ºW), in the oligotrophic central North Pacific (Inc 3, 4; 35 ºN, 145 ºW), and at Ocean Station PAPA (Inc 5, 6; 50 ºN, 145 ºW) in the subarctic North Pacific. For incubations 1-4, all incubation bottles were spiked with chelexed stocks of nitrate and phosphate, and aged (for trace metal cleanliness) silicic acid stocks, to target additions of 10 micromolar (µM) nitrate, silicic acid, and 0.8 µM phosphate; no macronutrients were added to Incs 5 and 6, which were already macronutrient replete. Replicates of pH treatment were additionally spiked with 1 nanomolar (nM) 57FeCl3 as a dissolved iron addition. Incubation bottles were labeled according to treatment and were the same across light bottles all incubations: A = pH 8.1, B = pH 8.1 + Fe, C = pH 7.6, D = pH 7.6 + Fe, E = pH 7.1, F = pH 7.1 + Fe. Replicates of each treatment were also incubated in heavy-duty black contractor bags to serve as dark controls (G = A, H = B, I = C, J = D, K = E, L = F), which were sampled on day final only.

Incubation sampling:
Triplicate bottles from each incubation were sampled daily over the course of the experiments. Incubation bottles were brought in from the incubators into a clean lab bubble in the ship, where they were washed down with Milli-Q and transferred into a clean hood. After gently inverting to mix, one liter of whole water was transferred into amber high-density polyethylene bottles for parallel filtering (< 100 millimeters (mm) Hg) of chlorophyll a on 5 µm membrane (Poretics) filters and on 0.7 µm GF/F (Whatman) filters and for filtering particulate organic carbon (POC), and particulate organic nitrogen (PON) filtering on combusted GF/F filters using a glass and stainless steel Millipore filtration rig in the main lab. Filters for chlorophyll a were frozen at -20 degrees Celsius (°C) in the dark prior to their extraction and analysis at sea; filters for POC and PON were wrapped in foil and stored in a -80 ºC freezer until analyzed on shore at the University of South Florida (MEC lab). The remaining contents of each bottle were filtered in the bubble clean hood on a custom acrylic filtration rig outfitted with dual stage Teflon filtration holders (Savillex) that allows the filtrate to go directly into sample bottles after passing through consecutive 5 µm and 0.4 µm acid-cleaned polycarbonate track-etched (PCTE; Whatman) filters. Samples for dissolved macronutrients were collected into acid-cleaned and triple-rinsed 15-milliliter (mL) polycarbonate Falcon tubes and stored in zipper bags in the fridge until analyzed shipboard following recommended practices (Becker et al., 2020), typically within 24 hours of collection (Caitlyn Parente, Kristen Buck lab). Samples for dissolved trace metals were collected in acid-cleaned and triple-rinsed narrow mouth low density polyethylene bottles, acidified with 0.024 molar (M) ultrapure hydrochloric acid (to pH ~1.8), and stored for shore-based analysis at the University of Nagasaki (Yoshiko Kondo). Samples for dissolved iron and nickel speciation were collected in acid-cleaned, Milli-Q-conditioned, and triple-rinsed narrow mouth fluorinated high density polyethylene bottles (Nalgene) and analyzed shipboard for dissolved iron speciation (Lise Artigue, Kristen Buck lab) before freezing at -20 ºC for shore-based dissolved nickel speciation analyses at Oregon State University (Matthew Koteskey, Kristen Buck lab). Filters containing the size-fractionated particulate material were folded into eighths, stored in acid-cleaned and dry snap-cap centrifuge tubes, and stored frozen at -20 ºC for shore-based particulate metal analyses at Oregon State University.

Sample analyses - macronutrients:
Filtered macronutrient samples were analyzed shipboard for phosphate, nitrate+nitrite, silicic acid, and nitrite on a QuAAtro39 AutoAnalyzer (SEAL Analytical) according to standard colorimetric methods (Strickland and Parsons, 1972). All reagents were prepared in dedicated labware with high purity Milli-Q (>18 MΩ cm) water. Working standards were prepared fresh daily in an artificial seawater (ASW; 35 grams per liter (g/L) sodium chloride, 0.5 g/L sodium bicarbonate) matrix using calibrated volumetric pipettes. Nine-point standard curves were analyzed at the beginning of each run with multiple reagent blanks. Quality control checks were analyzed every twelfth sample with ASW blanks and standards. The highest standard from the calibration curve was analyzed approximately every twenty samples to check for drift during the runs. Subsamples of reference material for nutrients in seawater (Konso) were measured in each run. Detection limits for each parameter were determined from three times the standard deviation of replicate lowest standards. Average limits of detection across the cruise dataset were 0.022 µM for phosphate, 0.108 µM for nitrate+nitrite, 0.107 µM for silicate, and 0.013 µM for nitrite. Values below these limits of detection are reported as 0 µM with accompanying QC Flag 6. Sample analyses for macronutrients were performed by MS student Caitlyn Parente shipboard.

Sample analyses - chlorophyll a:
Samples for chlorophyll a were placed in glass test tubes and 8 mL of 100% ethanol was added to each tube (Jespersen and Christoffersen, 1987; Wasmund et al., 2006). The tubes were capped and placed in the dark for the extraction at room temperature. After 12 hours, the fluorescence readings were subsequently measured following the standard acidification protocol (Parsons et al., 1984; Arar and Collins, 1992) using a Turner Designs model 10-AU fluorometer calibrated at the beginning of the cruise with pure chlorophyll a standards (Turner Designs; Anacystis nidulans) following standard JGOFS protocols (Knap et al., 1996).

Sample analyses – particulate organic carbon and nitrogen:
Combusted filters for POC and PON were dried in an oven at 450 ºC for 5 hours. Nitrogen and carbon isotope and bulk composition on the filters were measured by CF-EA-irms (Continuous Flow Elemental Analyzer Isotope Ratio Mass Spectrometry) at the University of South Florida College of Marine Science Marine Environmental Chemistry Laboratory using commonly accepted procedures (Werner et al 1999). Isotope compositions were measured on a ThermoFinnigan Delta+XL IRMS, are reported in per mil (‰) notation and are scaled to VPDB (d13C) and AT-Air (d15N). Secondary reference materials (NIST 8574 d13C = +37.63 ± 0.10 ‰, d15N = +47.57 ± 0.22 ‰, N = 9.52%, C = 40.81%, C:N (molar) = 5.0; NIST 8573 d13C = -26.39 ± 0.09‰, d15N = -4.52 ± 0.12‰ N = 9.52%, C = 40.81%, C:N (molar) = 5.0) were used to normalize raw measurements to the VPDB (d13C) and AT-Air (d15N) scales (Werner et al 2001, Qi et al 2003, Coplen et al 2006) and to calibrate elemental N, C and C:N. Measurement uncertainties, expressed as ±1 standard deviation of n=82 measurements of a laboratory reference material (NIST1577b d13C = -21.69 ± 0.14‰, d15N = 7.83 ± 0.16‰, %N = 9.95 ± 0.48%, %C =48.04 ± 0.71%, C:N (molar) = 5.63 ± 0.27) were ±0.11‰ for d13C ±0.20‰ for d15N, ±1.52 %RSD for N, ±1.73 %RSD for C, and ±1.94 %RSD for C:N.

945375_v1_shipboard_incubations.csv