Abstract Owing to its accurate distance measured from water-maser motions, NGC 4258 is one of the most important anchors for calibrating the Cepheid period–luminosity relations (PLRs). We expand on previous efforts and carry out a new Cepheid search in this system using the Hubble Space Telescope. We discover and measure a sample of 669 Cepheids in four new and two archival NGC 4258 fields, doubling the number of known Cepheids in this galaxy and obtaining an absolute calibration of their optical PLRs. We determine a Wesenheit (W _VI ^HST) PLR of − 2.574 ( ± 0.034 ) − 3.294 ( ± 0.042 ) log P , consistent with an independent Large Magellanic Cloud (LMC) calibration at the level of 0.032 ± 0.044 mag in its zero-point, after accounting for a metallicity dependence of −0.20 ± 0.05 mag dex⁻¹. Our determination of the PLR slope also agrees with the LMC-based value within their uncertainties. We attempt to characterize the metallicity effect of Cepheid PLRs using only the NGC 4258 sample, but a relatively narrow span of abundances limits our sensitivity and yields a W _VI ^HST zero-point dependence of −0.07 ± 0.21 mag dex⁻¹. The Cepheid measurements presented in this study have been used as part of the data to derive the Hubble constant in a companion paper by the SH0ES team.