AbstractWe present 1 year of in situ proton transfer reaction mass spectrometer (PTR‐MS) measurements of isoprene and its oxidation products methyl vinyl ketone (MVK) and methacrolein (MACR) from a 244 m tall tower in the U.S. Upper Midwest, located at an ecological transition between isoprene‐emitting deciduous forest and predominantly non‐isoprene‐emitting agricultural landscapes. We find that anthropogenic interferences (or anthropogenic isoprene) contribute on average 22% of the PTR‐MS m/z 69 signal during summer daytime, whereas MVK + MACR interferences (m/z 71) are minor (7%). After removing these interferences, the observed isoprene and MVK + MACR abundances show pronounced seasonal cycles, reaching summertime maxima of >2500 pptv (1 h mean). The tall tower is impacted both by nearby and more distant regional isoprene sources, with daytime enhancements of isoprene (but little MVK + MACR) under southwest winds and enhancements of MVK + MACR (but little isoprene) at other times. We find that the GEOS‐Chem atmospheric model with the MEGANv2.1 (Model of Emissions of Gases and Aerosols from Nature version 2.1) biogenic inventory can reproduce the isoprene observations to within model uncertainty given improved land cover and temperature estimates. However, a 60% low model bias in MVK + MACR cannot be resolved, even across diverse model assumptions for NOx emissions, chemistry, atmospheric mixing, dry deposition, land cover, and potential measurement interferences. This implies that, while isoprene emissions in the immediate vicinity of the tall tower are adequately captured, they are underestimated across the broader region. We show that this region experiences a strong seasonal shift between VOC‐limited chemistry during the spring and fall and NOx‐limited or transitional chemistry during the summer, driven by the spatiotemporal distribution of isoprene emissions. Isoprene's role in causing these chemical shifts is likely underestimated due to the underprediction of its regional emissions.