Lydia Villa-Komaroff (interviewed by Nafisa Syed)

Interview conducted with an MIT alumna as part of the Margaret MacVicar Memorial AMITA (Association of MIT Alumnae) Oral History Project. The purpose of the project is to document the life histories of women graduates of the Massachusetts Institute of Technology

Specific, temporally regulated expression of the insulin-like growth factor II gene during muscle cell differentiation

We have compared the expression of insulin-like growth factor II (IGF-II) messenger RNA (mRNA) to the expression of other mRNAs encoding proteins known to play pivotal roles during the differentiation of continuously cultured, fusing muscle cell lines. These cell lines respond to changes in culture conditions by undergoing a well characterized alteration in gene expression which leads to a change in their phenotype from dividing, mononucleate myoblasts to fused, multinucleate myotubes. The hallmarks of this differentiation program include the induction of myogenic regulatory genes as well as the genes that encode the contractile proteins. We have found that the differentiation of these cells leads to the production of multiple IGF-II transcripts. In one of the cell lines studied, C2C12, IGF-II mRNA levels were rapidly induced during differentiation. Increases in IGF-II mRNA levels preceded the expression of the contractile protein genes but occurred only after the activation of the myogenic regulatory gene myogenin. The same regulated pattern of IGF-II mRNA expression was seen in both rapidly and slowly fusing subclones of this cell line, indicating a requirement for IGF-II at a specific point during muscle differentiation. These results suggest that IGF-II plays an important role during the terminal differentiation of skeletal muscle cells and are consistent with the existence of an autocrine loop through which IGF-II may act to regulate the differentiation process

Characterization of the two nonallelic genes encoding mouse preproinsulin

We have cloned and sequenced the two mouse preproinsulin genes. The deduced amino acid sequences of the mature mouse insulins are identical to the published protein sequences. However, the nucleotide sequence indicates that the mouse I C-peptide has a deletion of two amino acids compared with the mouse II C-peptide. We used an S1 nuclease assay to confirm the presence of the deletion and to measure the ratio of transcripts from gene I to transcripts from gene II. The mouse preproinsulin I gene, like the rat gene I, is missing the second intervening sequence that normally interrupts the C-peptide region in other insulin genes. Comparison of the 5\u27 flanking sequences of the mouse and rat genes II indicates that they are homologous for at least 1000 base pairs. The preproinsulin I genes also share homology in their 5\u27 flanking DNAs; however, their homology to the preproinsulin II genes extends for only about 500 base pairs

Alternative 5\u27 exons either provide or deny an initiator methionine codon to the same alpha-tubulin coding region

The primary structures of two overlapping novel alpha-tubulin cDNA clones isolated from a Macaca fascicularis testis cDNA library and the corresponding human gene are presented. Although the general structure of the human gene conforms to that of previously described mammalian alpha-tubulin genes, there is a surprising difference: the ATG initiator codon is conspicuously absent. The macaque testis cDNA similarly lacks the initiator methionine, but otherwise encodes a variant alpha-tubulin isotype precisely conserved in the human gene. RNA blot analysis in the macaque, using a 3\u27 untranslated region probe, revealed the existence of two additional related transcripts expressed in every tissue examined except the adult testis. Sequence comparisons indicate that the 2.0 kb testis transcript and one of the additional transcripts result from differential transcription of the same gene. The two transcripts differ only at the 5\u27 end as a result of the recruitment of different 5\u27 exons. Curiously, the 5\u27 exon utilized outside the testis encodes an initiator methionine in the expected location

The ratio of mouse insulin I:insulin II does not reflect that of the corresponding preproinsulin mRNAs

Rats and mice both express two, non-allelic, insulin genes. In the rat the ratio of the two preproinsulin mRNAs closely matches that of the mature insulin peptides. The experiments reported here demonstrate that this is not the case in the mouse. The relative amounts of the two murine proinsulin RNAs were measured by an S1 nuclease assay. The ratio of preproinsulin I mRNA to preproinsulin II mRNA was 4:1 in RNA extracted from the pancreas of mice fed ad libitum or fasted for 72 h. A similar value was found in mouse islets of Langerhans after maintenance in tissue culture for 48 h at either 2.8 or 16.7 mM glucose. The ratio of insulin I:insulin II peptides, assessed by separating the two insulins using reversed phase high-performance liquid chromatography, was approximately 1:3 in both pancreas and islets. Thus in the mouse, unlike the rat, the ratio of the two insulin peptides does not reflect that of the two preproinsulin mRNAs