Reintroducing David Hoganis, a pioneer in molecular genetics and developmental biology. – Genes from the genome

Nowadays, we don’t think twice about running Q-PCR to check the expression of a gene of interest, or to sequence a genomic region to identify a mutation that causes an interesting phenotype. In contrast, 50 years ago, such a task could take an entire Ph.D. Molecular genetics has developed at a rapid pace, and its application to other fields has led to advances in our understanding of many physiological and pathological processes. David Hoganis, one of the pioneers of molecular genetics, developed several ground-breaking technologies for identifying genes and studying how their expression changes in different contexts. By linking technological advances with developmental biology and Drosophila Genetics, the Hoganis lab cloned key eukaryotic developmental genes and laid the foundation for genomic analysis in humans and many other species.^1-3

Despite using the technologies and molecular concepts pioneered by the Hoganis Lab on a daily basis, many current grad students and early career researchers in the field do not know the scientist who initially developed them. To celebrate Hogness ‘100^Th anniversary, and to teach the next generation of researchers the history behind certain concepts based on our genetic knowledge, the November issue of Genetics includes two back-to-back articles: one based on Hogans’ autobiography,² and one that explores the significance of an unpublished manuscript on ecdysone-mediated gene regulation that had languished on Hogan’s desk for more than 40 years.³

“Hogan was a true visionary and 10-15 years ahead of the field for these research questions. Drosophila”, say William Talbot and David Kingsley, co-authors of Current Perspectives in Genetics, which includes Hoganis’ autobiography.²

Take, for example, chromosomal walking, a method of loci cloning developed in the Hogans lab based on their genetic map locations. At that time (the mid-1970s), mutant alleles with interesting phenotypes were available for many different species and the genetic positions were known for some of them, but there was no way for researchers to link this information to the RNA or protein molecules produced by the gene. To solve this problem, the Hoganis lab digested genomic DNA and cloned the resulting fragments into bacterial vectors to generate a “library” of cloned genomic DNA fragments (plasmids, phage vectors, and cloning technology are currently in their infancy). These clones were then used to generate labeled probes that hybridized against the fly polytene chromosome to identify the exact genomic location of the clone’s DNA fragment. Starting with a clone around the mutation, the Hogans lab “walked” along the fly’s third chromosome by repeatedly isolating isolated clones. They used this strategy to identify and isolate clones containing well-known Betorex complex homeotic genes whose mutations alter the identity of body segments in Drosophila.⁴ This seminal work marked the molecular identification of the first key developmental genes in eukaryotes, and kick-started the era of molecular genetics.

Kingsley and Talbot, along with many others, were among the early adopters of chromosome walking technology. Convinced of the transformative power of this technology, they incorporated it into their research programs when starting their own labs and applied the method to different species (zebrafish for Talbot; Kingsley for mouse and stickleback fish). Tablebot, a former student in Hogans’ lab, and Kingsley are both in the development department at Stanford University, and Kingsley is both in the development department at Stanford University. After his office passed away and he had a trove of information, including handwritten conversations, grant applications, and some unpublished manuscripts. He carefully prepared these documents, which are now accessible through the Stanford Archives.

Among those documents, he found Hogans’ acceptance essay for Stanford’s inaugural Munzer Chair and realized it was the closest thing to an autobiography. Many good retreats and travesties have been written to highlight Dave’s exploits,^5-6 But what struck us was Dave’s own perspective,” says Talbot. Talbot and Kingsley said they wanted to share the autobiography with the wider scientific community, so others could read Dave Hogans’ own interpretation of his work. The team added some context to the document and the resulting perspective is now published in Genetics.²

Kingsley noted, “Hoggins was slow to write, but what he wrote was inspiring and passionate. While it was written decades ago, I found that my research could still shape today.” Hogans was widely known for his collaborative, polite nature, his remarkable training record, and his moderate publication rate. Pioneer shared his group’s key findings at conferences and seminars, and always made sure to credit any trainees or group members who contributed to the work. He was a strong advocate for his trainees and helped many of them set up their own labs. However, a scientist’s perfectionism can get the better of him when it comes to writing up and publishing his findings, and manuscripts can spend months to years on his desk.

Mariana Wilfner’s PhD project was the focus of one of these ill-fated manuscripts. Wolfner, currently a professor of molecular biology and genetics at Cornell University, was a graduate student in the Hoganis lab, where she worked with several other Hoganis lab members to identify and study ecdysone-mediated gene expression changes during larval migration. To identify which genes are regulated by ecdysone and study the molecular processes underlying metamorphosis, the team developed a different cDNA hybridization technology, which became a precursor to microarray technology. The scientists isolated salivary gland RNA from developmental stage animals and copied it into cDNA, which they cloned into plasmids to produce stage-specific libraries. They then prepared grids of individual cDNA clones from these staged libraries and blotted the clones’ DNA to a set of nitrocellulose membranes. Next, they exposed these membranes to a specific developmental stage with RNA-derived radiolabeled cDNA probes to identify clones of genes expressed at that time point. In parallel, they used radiolabeled cDNA from these clones to map the gene to the chromosome by in situ hybridization. “We were doing things that no one had ever done before, but there was this general sense of excitement in the lab and everyone believed that these experiments would work,” remembers Wolfner.

By combining these methods, Wolfner and colleagues showed how salivary gland RNA populations change during the larval-to-pupal transition, demonstrating the presence of robust gene regulatory mechanisms during development.³ Additionally, they identified and isolated specific genes whose expression was activated or repressed in response to ecdysone, which helped define the molecular pathways that underlie the developmental timing of this transition.

The team wrote a paper describing their work, but when Wolfner moved on to start his postdoc in Bruce Baker’s lab at UCSD, the final version of the paper remained in the Hogans queue, with copies bumping off Wolfner’s desk and those of his co-authors. Linda Risfo, now a professor of neurology at the University of Arizona, built on Wolfner’s work during graduate studies in the lab of Greg Guild, a former Hogan postdoc and co-author on Ecdyson’s manuscript. Restifo recently rediscovered the manuscript in its final form. Wanting to do justice to the seminal work and fill in the missing historical piece of the Ecdysone puzzle, she reached out to Wolfner. The pair teamed up with the remaining surviving original co-authors and Mark Griffinkel, the scientific grandson of Dave Hagins, such as Restofo, to review the discovery of ecdysone-related genes and the ecdysone signaling cascade. As an added bonus, he included the unpublished original manuscript as an appendix to his approach.³

Hogans’ work and findings revealed the potential of gene identification and unlocking scientists’ ability to study how gene expression changes over time and space in physiological and pathological contexts. Two papers published in the current issue of Genetics highlight how transformative Hogans’ work has been for the fields of molecular genetics and developmental biology, and will ensure that the next generation of researchers can rediscover this pioneer’s life’s work.