Keeping Track of the Genealogy of Heterozygotes Using Epigenetic Reference Codes and Breeding Tables

Abstract

Studying neurobehavioral consequences of the hypofunctional dopamine transporter (DAT) across several generations entails the need to monitor allelic transmission to offspring, taking into account both maternal and paternal inheritance. Since each type of heterozygote expresses differential phenotypes, based on lineage of inheritance for wild and mutated alleles (from male or female ancestors), it is important to track transgenerational epigenetic effects. We deemed it essential to assign specific abbreviations identifying their characteristics. Therefore, we devised a Mendelian-inspired table to keep track of these. Starting from two progenitors (WT and KO) we named resulting heterozygous progenies MAT and PAT to differentiate them based on inheritance of the wild allele (from the mother or father). Tracing subsequent generations, similar logic has been followed: if coupling HET dams with KO males, initials “M” [(grand)maternal] and “P” [(grand)paternal] are kept, but “AT” is turned into “IX” (MIX and PIX), while if breeding HETs with WTs, “M” is changed to “W” resembling an upside down “M” and “P” to “S” for “sperm” (WAT and SOT). To underline the development within “hyperdopaminergic-uterus” a central letter “U” is added (MUX, PUX, and QULL), while a Greek initial (μAT, μIX, and νIX) underlines the uterine-worsened origin of the allele. In HET × HET breeding (GIX and DIX), the mutated allele can be inherited from both sides of the genealogical line. However, when the mother is MAT, wild and mutated alleles encounter for the first time, causing putative anomalies in the progeny. Replacing dam with a second-generation female (MIX and MUX) may mitigate epigenetic effects on third-generation offspring; therefore suffixes (“-f,” “-fu,” “-ϕ,” and “-ϕu”) emphasize that subsequent-generation dams imply that the alleles already encountered in HET (rather than WT) grand-dams.

Truncated dopamine transporter's epigenetics: Heterozigosity of the grandmother rat temperates the vulnerable phenotype in second‐generation offspring

“…The first step was always dedicated to generating MAT pups from WT females bred with KO males. The next generation was obtained through the crossing of such atypical HET females, called MAT, again with a KO male, which gives rise to a progeny termed MIX rats (Liberati et al, 2022). The rats downstream of a MAT (thus, grandmother will be WT) can be divided into MIX‐by‐MAT (with mother MAT and father KO) or SOT (with a WT father); the rats downstream of a MIX (thus, grandmother will be HET) are MIX pups‐of‐a MIX, abbreviated to MIX 2 (with mother MIX and father KO) or SIX (with mother still MIX but a WT father), respectively.…”

“…Yet, to keep it simple, we focused on circadian cycles and sociability: These two endpoints are indeed known to be modulated by the paternal versus maternal lineage of inheritance for DAT (Oggiano et al, 2022). A full battery of tests, including the Plus‐Maze and Porsolt tests, is ongoing on other HET epigenotypes (for reference, see Liberati et al, 2022).…”

“…Third-generation "atypical" heterozygote; SIX, SOT-gestated-by-MIX, with MAT grand-dam, thus offspring of MIX dam  WT father. Third-generation "atypical" heterozygote, inverted asset compared with MIX 2 (Liberati et al, 2022); SOT, sperm-originated wild/healthy allele, thus offspring of a MAT dam  WT father. Second-generation "atypical" heterozygote, with inverted asset compared to MIX (Liberati et al, 2022).…”

“…Then, present aim was to assess the successive generations, produced by the MAT females used as mothers. As a first approach, we bred MAT dams with KO male breeders to obtain a female HET offspring that was called MIX, as representing a portion of the "classical, mixed" offspring (Liberati et al, 2022). Both MAT and MIX are HET subjects with wild/healthy allele inherited by the mother (i.e., "asset M").…”

“…Notably, this latter group of rats had now a HET grandmother, rather than a WT one. Being otherwise all asset-P HET subjects, with HET dams, the comparison of SOT vs. SIX rats allowed again the comparison between genotypes of the grandmother (see Liberati et al, 2022).…”